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Genome-Wide Expression Patterns in Saccharomyces cerevisiae: Comparison of Drug Treatments and Genetic Alterations Affecting Biosynthesis of Ergosterol

Bammert, Gary F.; Fostel, Jennifer M.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /05/2000 EN
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882.30945%
Enzymes in the ergosterol-biosynthetic pathway are the targets of a number of antifungal agents including azoles, allylamines, and morpholines. In order to understand the response of Saccharomyces cerevisiae to perturbations in the ergosterol pathway, genome-wide transcript profiles following exposure to a number of antifungal agents targeting ergosterol biosynthesis (clotrimazole, fluconazole, itraconazole, ketoconazole, voriconazole, terbinafine, and amorolfine) were obtained. These profiles were compared to the transcript profiles of strains containing deletions of one of the late-stage ergosterol genes: ERG2, ERG5, or ERG6. A total of 234 genes were identified as responsive, including the majority of genes from the ergosterol pathway. Expression of several responsive genes, including ERG25, YER067W, and YNL300W, was also monitored by PCR over time following exposure to ketoconazole. The kinetics of transcriptional response support the conditions selected for the microarray experiment. In addition to ergosterol-biosynthetic genes, 36 mitochondrial genes and a number of other genes with roles related to ergosterol function were responsive, as were a number of genes responsive to oxidative stress. Transcriptional changes related to heme biosynthesis were observed in cells treated with chemical agents...

Candidacidal Activities of Human Lactoferrin Peptides Derived from the N Terminus

Lupetti, Antonella; Paulusma-Annema, Akke; Welling, Mick M.; Senesi, Sonia; van Dissel, Jaap T.; Nibbering, Peter H.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /12/2000 EN
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882.2475%
In light of the need for new antifungal agents, the candidacidal activities of human lactoferrin (hLF) and synthetic peptides representing the first, hLF(1-11), and second, hLF(21-31), cationic domains of its N terminus were compared. The results revealed that hLF(1-11) was more effective in killing fluconazole-resistant Candida albicans than hLF(21-31) and much more effective than lactoferrin, as determined microbiologically and by propidium iodide (PI) staining. By using hLF(1-11) and various derivatives, it was found that the second and third residues of the N terminus of hLF(1-11) were critical for its candidacidal activity. Detailed investigation to elucidate the mechanism of action of hLF(1-11) revealed a dose-dependent release of ATP by Candida upon exposure to hLF(1-11). Our observations that sodium azide reduced the PI uptake and candidacidal activity of hLF(1-11) and that, upon exposure to hLF(1-11), the fluorescent dye rhodamine 123 first accumulated inside the mitochondria and later was released into the cytoplasm indicate that the peptide triggers the energized mitochondrion. Furthermore, oxidized ATP, which interferes with the interaction of ATP with its extracellular receptors, blocked the candidacidal action of hLF(1-11)...

Sordarins: A New Class of Antifungals with Selective Inhibition of the Protein Synthesis Elongation Cycle in Yeasts

Domínguez, Juan Manuel; Kelly, Valerie A.; Kinsman, Oonagh S.; Marriott, Michael S.; Gómez de las Heras, Federico; Martín, J. Julio
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /09/1998 EN
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883.3749%
GR135402, a sordarin derivative, was isolated in an antifungal screening program. GR135402, sordarin, and derivatives of both compounds were evaluated for their ability to inhibit cell-free translational systems from five different pathogenic fungi (Candida albicans, Candida glabrata, Candida krusei, Candida parapsilosis, and Cryptococcus neoformans). The activity profile of GR135402 is extended to other chemical compounds derived from sordarin. Experimental results indicate that sordarin analogs exert their antifungal effects by specifically inhibiting the protein synthesis elongation cycle in yeasts but do not affect protein synthesis machinery in mammalian systems. Intrinsically resistant strains owe their resistance to differences in the molecular target of sordarins in these strains. Preliminary studies performed to elucidate the mode of action of this new class of antifungal agents have shown that the putative target of sordarins is one of the protein synthesis elongation factors.

Transcriptional Profile of the Escherichia coli Response to the Antimicrobial Insect Peptide Cecropin A

Hong, Robert W.; Shchepetov, Mikhail; Weiser, Jeffrey N.; Axelsen, Paul H.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /01/2003 EN
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883.2908%
Cationic antimicrobial peptides are believed to exert their primary activities on anionic bacterial cell membranes; however, this model does not adequately account for several important structure-activity relationships. These relationships are likely to be influenced by the bacterial response to peptide challenge. In order to characterize the genomic aspect of this response, transcription profiles were examined for Escherichia coli isolates treated with sublethal and lethal concentrations of the cationic antimicrobial peptide cecropin A. Transcript levels for 26 genes changed significantly following treatment with sublethal peptide concentrations, and half of the transcripts corresponded to protein products with unknown function. The pattern of response is distinct from that following treatment with lethal concentrations and is also distinct from the bacterial response to nutritional, thermal, osmotic, or oxidative stress. These results demonstrate that cecropin A induces a genomic response in E. coli apart from any lethal effects on the membrane and suggest that a complete understanding of its mechanism of action may require a detailed examination of this response.

In Vitro Bactericidal Activities of Daptomycin against Staphylococcus aureus and Enterococcus faecalis Are Not Mediated by Inhibition of Lipoteichoic Acid Biosynthesis

Laganas, Valerie; Alder, Jeffrey; Silverman, Jared A.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /08/2003 EN
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882.0219%
Previous studies have suggested that lipoteichoic acid biosynthesis inhibition is the mechanism of action of daptomycin. In this investigation, daptomycin inhibited all macromolecular synthesis in Staphylococcus aureus, Enterococcus faecalis, and Enterococcus hirae without kinetic or dose specificity for lipoteichoic acid. Daptomycin remained bactericidal in the absence of ongoing lipoteichoic acid synthesis. Inhibition of lipoteichoic acid synthesis is apparently not the mechanism of action of daptomycin in these pathogens.

Possible Mechanism of Miltefosine-Mediated Death of Leishmania donovani†

Verma, Navin K.; Dey, Chinmoy S.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /08/2004 EN
Relevância na Pesquisa
882.0219%
Miltefosine causes leishmanial death, but the possible mechanism(s) of action is not known. The mode of action of miltefosine was investigated in vitro in Leishmania donovani promastigotes as well as in extra- and intracellular amastigotes. Here, we demonstrate that miltefosine induces apoptosis-like death in L. donovani based on observed phenomena such as nuclear DNA condensation, DNA fragmentation with accompanying ladder formation, and in situ labeling of DNA fragments by the terminal deoxyribonucleotidyltransferase-mediated dUTP-biotin nick end labeling method. Understanding of miltefosine-mediated death will facilitate the design of new therapeutic strategies against Leishmania parasites.

Sodium Antimony Gluconate Induces Generation of Reactive Oxygen Species and Nitric Oxide via Phosphoinositide 3-Kinase and Mitogen-Activated Protein Kinase Activation in Leishmania donovani-Infected Macrophages

Mookerjee Basu, Jayati; Mookerjee, Ananda; Sen, Prosenjit; Bhaumik, Suniti; Sen, Pradip; Banerjee, Subha; Naskar, Ksudiram; Choudhuri, Soumitra K.; Saha, Bhaskar; Raha, Sanghamitra; Roy, Syamal
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /05/2006 EN
Relevância na Pesquisa
882.21195%
Pentavalent antimony complexes, such as sodium stibogluconate and sodium antimony gluconate (SAG), are still the first choice for chemotherapy against various forms of leishmaniasis, including visceral leishmaniasis, or kala-azar. Although the requirement of a somewhat functional immune system for the antileishmanial action of antimony was reported previously, the cellular and molecular mechanism of action of SAG was not clear. Herein, we show that SAG induces extracellular signal-regulated kinase 1 (ERK-1) and ERK-2 phosphorylation through phosphoinositide 3-kinase (PI3K), protein kinase C, and Ras activation and p38 mitogen-activated protein kinase (MAPK) phosphorylation through PI3K and Akt activation. ERK-1 and ERK-2 activation results in an increase in the production of reactive oxygen species (ROS) 3 to 6 h after SAG treatment, while p38 MAPK activation and subsequent tumor necrosis factor alpha release result in the production of nitric oxide (NO) 24 h after SAG treatment. Thus, this study has provided the first evidence that SAG treatment induces activation of some important components of the intracellular signaling pathway, which results in an early wave of ROS-dependent parasite killing and a stronger late wave of NO-dependent parasite killing. This opens up the possibility of this metalloid chelate being used in the treatment of various diseases either alone or in combination with other drugs and vaccines.

New Azasterols against Trypanosoma brucei: Role of 24-Sterol Methyltransferase in Inhibitor Action

Gros, Ludovic; Castillo-Acosta, Victor Manuel; Jiménez, Carmen Jiménez; Sealey-Cardona, Marco; Vargas, Sofia; Manuel Estévez, Antonio; Yardley, Vanessa; Rattray, Lauren; Croft, Simon L.; Ruiz-Perez, Luis M.; Urbina, Julio A.; Gilbert, Ian H.; Pacanowsk
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /08/2006 EN
Relevância na Pesquisa
882.1665%
A series of azasterol derivatives, designed as potential inhibitors of the Δ24-sterol methyltransferase enzyme (24-SMT), were synthesized and evaluated for their activities against parasitic protozoa. Values in the nanomolar range were obtained for 50% effective dose against the Trypanosoma brucei subsp. rhodesiense bloodstream form cultured in vitro. In order to investigate the mode of action, Trypanosoma brucei subsp. brucei 24-SMT was cloned and overexpressed and compounds were assayed for inhibitory activity. None of the inhibitors tested appeared to be active against the enzyme. Sterol composition analysis showed that only cholestane type sterols are present in membranes of bloodstream forms while ergosterol is a major component of procyclic sterol extracts. Interestingly, Northern blot analysis showed the presence of 24-SMT mRNA in both the procyclic and the bloodstream forms of the parasite, although levels of mRNA were threefold lower in the latter. Likewise, Western blot analysis and activity determinations evidenced the existence of active enzyme in both forms of the parasite. We conclude that the designed compounds act at sites other than 24-SMT in Trypanosoma brucei.

Effect of PEX, a Noncatalytic Metalloproteinase Fragment with Integrin-Binding Activity, on Experimental Chlamydophila pneumoniae Infection

Caronzolo, Dario; Lucini, Valeria; Pannacci, Marilou; Grosso, Silvia; Kieffer, Nelly; Bello, Lorenzo; Bikfalvi, Andreas; Scaglione, Francesco
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /10/2006 EN
Relevância na Pesquisa
883.2908%
Chlamydophila pneumoniae is a pathogen that is involved in acute and chronic respiratory infections and that is associated with asthma and coronary artery diseases. In this study, we evaluated the effects of PEX, a noncatalytic metalloproteinase fragment with integrin-binding activity, against experimental infections caused by C. pneumoniae. Moreover, we investigated the relationships between C. pneumoniae and αvβ3 integrin functions in order to explain the possible mechanism of action of PEX both in vitro and in vivo. For the in vitro experiments, HeLa cells were infected with C. pneumoniae and treated with either PEX or azithromycin. The results obtained with PEX were not significantly different (P > 0.05) from those achieved with azithromycin. Similar results were also obtained in a lung infection model. Male C57BL/J6 mice inoculated intranasally with 106 inclusion-forming units of C. pneumoniae were treated with either PEX or azithromycin plus rifampin. Infected mice treated with PEX showed a marked decrease in C. pneumoniae counts versus those for the controls; this finding did not differ significantly (P > 0.05) from the results observed for the antibiotic-treated group. Integrin αvβ3 plays an important role in C. pneumoniae infection. Blockage of integrin activation led to a significant inhibition of C. pneumoniae infection in HeLa cells. Moreover...

Azurin-Like Protein Blocks Invasion of Toxoplasma gondii through Potential Interactions with Parasite Surface Antigen SAG1▿

Naguleswaran, Arunasalam; Fialho, Arsenio M.; Chaudhari, Anita; Hong, Chang Soo; Chakrabarty, Ananda M.; Sullivan, William J.
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
EN
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883.2908%
Some pathogenic bacteria produce factors that have evolved a capacity to neutralize competing microbes. The cupredoxin family protein azurin, produced by Pseudomonas aeruginosa, exhibits a remarkable ability to impede invasion of a number of diverse intracellular pathogens, including the human AIDS virus human immunodeficiency virus type 1 and the protozoan parasite Plasmodium falciparum (which causes malaria). Here we report that azurin and an azurin-like protein (Laz) from gonococci/meningococci have activity against Toxoplasma, an apicomplexan parasite that causes opportunistic infection in immunocompromised individuals. We demonstrate that the mechanism of action for Laz involves interfering with the ability of Toxoplasma to adhere to host cells. Computer structural analysis reveals that azurin shares structural features with the predominant surface antigen SAG1, which is known to play an important role in parasite attachment. Interestingly, azurin also has structural similarities to a monoclonal antibody to SAG1. Surface plasmon resonance binding studies validate that SAG1 interacts strongly with Laz and, to lesser extent, azurin. Moreover, Toxoplasma mutants lacking SAG1 are not as susceptible to the growth-inhibitory effects of Laz. Collectively...

Modulation of Gene Expression in Human Macrophages Treated with the Anti-Leishmania Pentavalent Antimonial Drug Sodium Stibogluconate▿ †

El Fadili, Karima; Imbeault, Michaël; Messier, Nadine; Roy, Gaétan; Gourbal, Benjamin; Bergeron, Marc; Tremblay, Michel J.; Légaré, Danielle; Ouellette, Marc
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
EN
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882.2308%
Within the mammalian host, Leishmania donovani is an obligatory intracellular protozoan parasite that resides and multiplies exclusively in the phagolysosomes of macrophages. Leishmania control relies primarily on chemotherapy, with the mainstay being pentavalent antimony (SbV) complexed to carbohydrates in the form of sodium stibogluconate (Pentostam) or meglumine antimoniate (Glucantime). The mode of action of SbV is still not known precisely. To explore the effect of SbV on macrophage gene expression, a microarray analysis was performed using Affymetrix focus arrays to compare gene expression profiles in noninfected and L. donovani-infected THP-1 monocytic cells treated or not treated with sodium stibogluconate. Under our experimental conditions, SbV changed the expression of a few host genes, and this was independent of whether cells were infected or not infected with Leishmania. Leishmania infection had a greater effect on the modulation of host gene expression. Statistical analyses have indicated that the expression of eight genes was modified by at least twofold upon SbV treatment, with six genes upregulated and two genes downregulated. One gene whose expression was affected by SbV was the heme oxygenase gene HMOX-1, and this change was observed both in the monocytic cell line THP-1 and in primary human monocyte-derived macrophages. Another pathway that was affected was the glutathione biosynthesis pathway...

Praziquantel Affects the Regulatory Myosin Light Chain of Schistosoma mansoni▿

Gnanasekar, Munirathinam; Salunkhe, Ashok M.; Mallia, A. Krishna; He, Yi Xun; Kalyanasundaram, Ramaswamy
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
882.1665%
Praziquantel (PZQ) is the drug of choice for schistosomiasis and probably is the only highly effective drug currently available for treating schistosomiasis-infected individuals. The mode of action of PZQ involves increasing the calcium uptake of the parasite, resulting in tegumental damage and death of the parasite. Despite its remarkable function, the target of PZQ has not been identified yet. To begin to understand where PZQ acts, in this study we expressed the cDNA library of Schistosoma mansoni on the surface of T7 bacteriophages and screened this library with labeled PZQ. This procedure identified a clone that strongly bound to PZQ. Subsequent DNA analysis of inserts showed that the clone coded for regulatory myosin light chain protein. The gene was then cloned, and recombinant S. mansoni myosin light chain (SmMLC) was expressed. Immunoblot analysis using antibodies raised to recombinant SmMLC (rSmMLC) showed that SmMLC is abundantly expressed in schistosomula and adult stages compared to the amount in cercarial stages. In vitro analyses also confirmed that PZQ strongly binds to rSmMLC. Further, peptide mapping studies showed that PZQ binds to amino acids 46 to 76 of SmMLC. Immunoprecipitation analysis confirmed that SmMLC is phosphorylated in vivo upon exposure to PZQ. Interestingly...

Antifungal Activity of Tamoxifen: In Vitro and In Vivo Activities and Mechanistic Characterization▿ †

Dolan, Kristy; Montgomery, Sara; Buchheit, Bradley; DiDone, Louis; Wellington, Melanie; Krysan, Damian J.
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
882.1385%
Tamoxifen (TAM), an estrogen receptor antagonist used primarily to treat breast cancer, has well-recognized antifungal properties, but the activity of TAM has not been fully characterized using standardized (i.e., CLSI) in vitro susceptibility testing, nor has it been demonstrated in an in vivo model of fungal infection. In addition, its mechanism of action remains to be clearly defined at the molecular level. Here, we report that TAM displays in vitro activity (MIC, 8 to 64 μg/ml) against pathogenic yeasts (Candida albicans, other Candida spp., and Cryptococcus neoformans). In vivo, 200 mg/kg of body weight per day TAM reduced kidney fungal burden (−1.5 log10 CFU per g tissue; P = 0.008) in a murine model of disseminated candidiasis. TAM is a known inhibitor of mammalian calmodulin, and TAM-treated yeast show phenotypes consistent with decreased calmodulin function, including lysis, decreased new bud formation, disrupted actin polarization, and decreased germ tube formation. The overexpression of calmodulin suppresses TAM toxicity, hypofunctional calmodulin mutants are hypersensitive to TAM, and TAM interferes with the interaction between Myo2p and calmodulin, suggesting that TAM targets calmodulin as part of its mechanism of action. Taken together...

Mode-of-Action Studies of the Novel Bisquaternary Bisnaphthalimide MT02 against Staphylococcus aureus▿ †

Menzel, Thomas M.; Tischer, Maximilian; François, Patrice; Nickel, Joachim; Schrenzel, Jacques; Bruhn, Heike; Albrecht, Annette; Lehmann, Leane; Holzgrabe, Ulrike; Ohlsen, Knut
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
EN
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883.2908%
Screening of various bisquaternary bisnaphthalimides against a variety of human pathogens revealed one compound, designated MT02, with strong inhibitory effects against Gram-positive bacteria. The MICs ranged from 0.31 μg/ml against community-acquired methicillin-resistant Staphylococcus aureus (MRSA) lineage USA300 to 20 μg/ml against Streptococcus pneumoniae. Radioactive whole-cell labeling experiments indicated a strong impact of MT02 on bacterial DNA replication. DNA microarray studies generated a transcriptional signature characterized by stronger expression of genes involved in DNA metabolism, DNA replication, SOS response, and transport of positively charged compounds. Furthermore, surface plasmon resonance and gel retardation experiments demonstrated direct binding of MT02 to DNA in a concentration-dependent, reversible, and non-sequence-specific manner. The data presented suggest that the bisquaternary bisnaphthalimide MT02 exerts anti-Gram-positive activity by binding to DNA and thereby preventing appropriate DNA replication.

The In Vitro Contribution of Autolysins to Bacterial Killing Elicited by Amoxicillin Increases with Inoculum Size in Enterococcus faecalis▿ †

Dubée, Vincent; Chau, Françoise; Arthur, Michel; Garry, Louis; Benadda, Samira; Mesnage, Stéphane; Lefort, Agnès; Fantin, Bruno
Fonte: American Society for Microbiology (ASM) Publicador: American Society for Microbiology (ASM)
Tipo: Artigo de Revista Científica
EN
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881.0608%
The mechanisms of antibiotic-induced cell death are poorly understood despite the critical role of the bactericidal activities of antibiotics for successful treatment of severe infections. These mechanisms include irreversible damaging of macromolecules by reactive oxygen species and bacteriolysis mediated by peptidoglycan hydrolases (autolysins). We have assessed the contribution of the second mechanism by using an autolysin-deficient mutant of Enterococcus faecalis and shown that it contributes to amoxicillin-induced cell lysis only at a high bacterial density.

Bioactivity and the First Transmission Electron Microscopy Immunogold Studies of Short De Novo-Designed Antimicrobial Peptides▿

Azad, Marisa Ann; Huttunen-Hennelly, Heidi Esther Katrina; Ross Friedman, Cynthia
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /05/2011 EN
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882.0219%
In light of the era of microbial drug resistance, the current study aimed to better understand the relationships between sequence, higher-order structure, and mechanism of action for five designed peptides against multidrug-resistant (MDR) pathogens. All peptides studied were 15 residues long, were polycationic, adopted alpha-helical structures within hydrophobic environments (excluding the d-amino acid-substituted peptide MA-d), and contained N-terminal glycine residues, a novel antimicrobial peptide (AMP) design principle. Increasing hydrophobicity enhanced MICs (≤500 μg/ml to ≤7.4 μg/ml) without significantly increasing hemolytic activity (18% maximum hemolysis at 3,400 μg/ml). To the best of our knowledge, this is the first study to have successfully adapted and used a transmission electron microscopy (TEM) immunogold method to investigate the mechanism of action of short (∼15 residues long) AMPs within bacteria. We propose a “floodgate” mechanism to possibly explain membrane deformation and the relative absence of membrane-associated peptides 10 h into incubation.

Effects of Antimicrobial Peptides on Methanogenic Archaea

Bang, C.; Schilhabel, A.; Weidenbach, K.; Kopp, A.; Goldmann, T.; Gutsmann, T.; Schmitz, R. A.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /08/2012 EN
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881.8746%
As members of the indigenous human microbiota found on several mucosal tissues, Methanobrevibacter smithii and Methanosphaera stadtmanae are exposed to the effects of antimicrobial peptides (AMPs) secreted by these epithelia. Although antimicrobial and molecular effects of AMPs on bacteria are well described, data for archaea are not available yet. Besides, it is not clear whether AMPs affect them as the archaeal cell envelope differs profoundly in terms of chemical composition and structure from that of bacteria. The effects of different synthetic AMPs on growth of M. smithii, M. stadtmanae, and Methanosarcina mazei were tested using a microtiter plate assay adapted to their anaerobic growth requirements. All three tested methanoarchaea were highly sensitive against derivatives of human cathelicidin, of porcine lysin, and a synthetic antilipopolysaccharide peptide (Lpep); however, sensitivities differed markedly among the methanoarchaeal strains. The potent AMP concentrations affecting growth were below 10 μM, whereas growth of Escherichia coli WBB01 was not affected at peptide concentrations up to 10 μM under the same anaerobic growth conditions. Atomic force microscopy and transmission electron microscopy revealed that the structural integrity of the methanoarchaeal cells is destroyed within 4 h after incubation with AMPs. The disruption of the cell envelope of M. smithii...

Insights into the Mode of Action of Novel Fluoroketolides, Potent Inhibitors of Bacterial Protein Synthesis

Krokidis, Marios G.; Márquez, Viter; Wilson, Daniel N.; Kalpaxis, Dimitrios L.; Dinos, George P.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /01/2014 EN
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883.3749%
Ketolides, the third generation of expanded-spectrum macrolides, have in the last years become a successful weapon in the endless war against macrolide-resistant pathogens. Ketolides are semisynthetic derivatives of the naturally produced macrolide erythromycin, displaying not only improved activity against some erythromycin-resistant strains but also increased bactericidal activity as well as inhibitory effects at lower drug concentrations. In this study, we present a series of novel ketolides carrying alkyl-aryl side chains at the C-6 position of the lactone ring and, additionally, one or two fluorine atoms attached either directly to the lactone ring at the C-2 position or indirectly via the C-13 position. According to our genetic and biochemical studies, these novel ketolides occupy the known macrolide binding site at the entrance of the ribosomal tunnel and exhibit lower MIC values against wild-type or mutant strains than erythromycin. In most cases, the ketolides display activities comparable to or better than the clinically used ketolide telithromycin. Chemical protection experiments using Escherichia coli ribosomes bearing U2609C or U754A mutations in 23S rRNA suggest that the alkyl-aryl side chain establishes an interaction with the U2609-A752 base pair...

Antiparasitic Effect of Vitamin B12 on Trypanosoma cruzi

Ciccarelli, Alejandra B.; Frank, Fernanda M.; Puente, Vanesa; Malchiodi, Emilio L.; Batlle, Alcira; Lombardo, Maria Elisa
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /10/2012 EN
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882.2475%
A nutritional characteristic of trypanosomatid protozoa is that they need a heme compound as a growth factor. Because of the cytotoxic activity of heme and its structural similarity to cobalamins, we have investigated the in vitro and in vivo effect of vitamin B12 (or cyanocobalamin) on the different forms of Trypanosoma cruzi. Cyanocobalamin showed a marked antiparasitic activity against epimastigotes (50% inhibitory concentration [IC50], 2.42 μM), amastigotes (IC50, 10.69 μM), and trypomastigotes (IC50, 9.46 μM). Anti-epimastigote and -trypomastigote values were 1.7 to 4 times lower than those obtained with the reference drug benznidazole (Bnz). We also found that B12 and hemin do not interact with each other in their modes of action. Our results show that B12 increases intracellular oxidative activity and stimulates both superoxide dismutase (50%) and ascorbate peroxidase (20%) activities, while the activity of trypanothione reductase was not modified. In addition, we found that the antioxidants dithiothreitol and ascorbic acid increase the susceptibility of the parasite to the cytotoxic action of B12. We propose that vitamin B12 exerts its growth-inhibitory effect through the generation of reactive oxygen species. In an in vivo assay...

Correlation of Daptomycin Bactericidal Activity and Membrane Depolarization in Staphylococcus aureus

Silverman, Jared A.; Perlmutter, Nancy G.; Shapiro, Howard M.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: Artigo de Revista Científica
Publicado em /08/2003 EN
Relevância na Pesquisa
882.106%
The objective of this study was to further elucidate the role of membrane potential in the mechanism of action of daptomycin, a novel lipopeptide antibiotic. Membrane depolarization was measured by both fluorimetric and flow cytometric assays. Adding daptomycin (5 μg/ml) to Staphylococcus aureus gradually dissipated membrane potential. In both assays, cell viability was reduced by >99% and membrane potential was reduced by >90% within 30 min of adding daptomycin. Cell viability decreased in parallel with changes in membrane potential, demonstrating a temporal correlation between bactericidal activity and membrane depolarization. Decreases in viability and potential also showed a dose-dependent correlation. Depolarization is indicative of ion movement across the cytoplasmic membrane. Fluorescent probes were used to demonstrate Ca2+-dependent, daptomycin-triggered potassium release from S. aureus. Potassium release was also correlated with bactericidal activity. This study demonstrates a clear correlation between dissipation of membrane potential and the bactericidal activity of daptomycin. A multistep model for daptomycin's mechanism of action is proposed.