RESUMEN

Methicillin-resistant Staphylococcus aureus (MRSA) is responsible for high morbidity and mortality rates. Citral has been studied in the pharmaceutical industry and has shown antimicrobial activity. This study aimed to analyze the antimicrobial activity of citral in inhibiting biofilm formation and modulating virulence genes, with the ultimate goal of finding a strategy for treating infections caused by MRSA strains. Citral showed antimicrobial activity against MRSA isolates with minimum inhibitory concentration (MIC) values between 5 mg/mL (0.5%) and 40 mg/mL (4%), and minimum bactericidal concentration (MBC) values between 10 mg/mL (1%) and 40 mg/mL (4%). The sub-inhibitory dose was 2.5 mg/mL (0.25%). Citral, in an antibiogram, modulated synergistically, antagonistically, or indifferent to the different antibiotics tested. Prior to evaluating the antibiofilm effects of citral, we classified the bacteria according to their biofilm production capacity. Citral showed greater efficacy in the initial stage, and there was a significant reduction in biofilm formation compared to the mature biofilm. qPCR was used to assess the modulation of virulence factor genes, and icaA underexpression was observed in isolates 20 and 48. For icaD, seg, and sei, an increase was observed in the expression of ATCC 33,591. No significant differences were found for eta and etb. Citral could be used as a supplement to conventional antibiotics for MRSA infections.

Asunto(s)

Monoterpenos Acíclicos/farmacología , Antibacterianos/farmacología , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Biopelículas/efectos de los fármacos , Staphylococcus aureus Resistente a Meticilina/patogenicidad , Pruebas de Sensibilidad Microbiana , Microscopía Confocal , Factores de Virulencia/antagonistas & inhibidores

RESUMEN

Identification of allosteric inhibitors of PTPs has attracted great interest as a new strategy to overcome the challenge of discover potent and selective molecules for therapeutic intervention. YopH is a virulence factor of the genus Yersinia, validated as an antimicrobial target. The finding of a second substrate binding site in YopH has revealed a putative allosteric site that could be further exploited. Novel chalcone compounds that inhibit PTPs activity were designed and synthesized. Compound 3j was the most potent inhibitor, interestingly, with different mechanisms of inhibition for the panel of enzymes evaluated. Further, our results showed that compound 3j is an irreversible non-competitive inhibitor of YopH that binds to a site different than the catalytic site, but close to the well-known second binding site of YopH.

Asunto(s)

Proteínas de la Membrana Bacteriana Externa/antagonistas & inhibidores , Chalcona/farmacología , Inhibidores Enzimáticos/farmacología , Proteínas Tirosina Fosfatasas/antagonistas & inhibidores , Factores de Virulencia/antagonistas & inhibidores , Sitio Alostérico/efectos de los fármacos , Proteínas de la Membrana Bacteriana Externa/metabolismo , Chalcona/síntesis química , Chalcona/química , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Estructura Molecular , Proteínas Tirosina Fosfatasas/metabolismo , Relación Estructura-Actividad , Factores de Virulencia/metabolismo

RESUMEN

Despite the main strategy to overcome bacterial resistance has focused on the development of more potent antimicrobial agents, the evolutionary pressure caused by such drugs makes this strategy limited. Molecules that interfere with virulence factors appear as a promising alternative though, as they cause reduced selective pressure. As a matter of fact, staphyloxanthin biosynthesis inhibition (STXBI) has been pursued as promising strategy to reduce S. aureus virulence. Herein, we report the inhibitory profile of 27 tetrangomycin derivatives over staphyloxanthin production. The experimental result showed that naphthoquinone dehydro-α-lapachone (25 - EC50 = 57.29 ± 1.15 µM) and 2-Isopropylnaphtho[2,3-b]furan-4,9-dione (26 EC50 = 82.10 ± 1.09 µM) are the most potent compounds and suggest that hydrogen acceptor groups and lipophilic moieties decorating the naphthoquinone ring are crucial for STXBI. In addition, we present an in situ analysis, through RAMAN spectroscopy, that is inexpensive and might be employed to probe the mechanism of action of staphyloxanthin biosynthesis inhibitors. Therefore, our molecular simplification strategies afforded promising lead compounds for the development of drugs that modulate S. aureus staphyloxanthin biosynthesis.

Asunto(s)

Antibacterianos/farmacología , Naftoquinonas/farmacología , Staphylococcus aureus/metabolismo , Xantófilas/metabolismo , Benzo(a)Antracenos/química , Benzo(a)Antracenos/farmacología , Farmacorresistencia Bacteriana Múltiple , Naftalenos/química , Naftalenos/farmacología , Naftoquinonas/química , Staphylococcus aureus/efectos de los fármacos , Relación Estructura-Actividad , Factores de Virulencia/antagonistas & inhibidores , Factores de Virulencia/biosíntesis

RESUMEN

Pseudomonas aeruginosa is the second most emerging multidrug-resistant, opportunistic pathogen after Acinetobacter baumannii that poses a threat in nursing homes, hospitals, and patients who need devices such as ventilators and blood catheters. Its ability to form quorum sensing-regulated virulence factors and biofilm makes it more resistant to top most therapeutic agents such as carbapenems and next-generation antibiotics. In the current study, we studied the quorum quenching potential of secondary metabolites of Mycoleptodiscus indicus PUTY1 strain. In vitro observation showed a mitigation in virulence factors such as rhamnolipids, protease, elastase pyocyanin, exopolysaccharides, and hydrogen cyanide gas. Furthermore, a significant reduction in the motility such as swimming, swarming, twitching, and inhibition in biofilm formation by Pseudomonas aeruginosa PAO1 was observed. Results of in vitro studies were further confirmed by in silico studies through docking and molecular dynamic simulation of GC-MS-detected compounds of Mycoleptodiscus indicus employing LasR and RhlR proteins. Both in vitro and in silico observations indicate a new alternative approach for combating virulence of Pseudomonas aeruginosa by targeting its protein receptors LasR and RhlR. Graphical abstract.

Asunto(s)

Ascomicetos/química , Biopelículas/efectos de los fármacos , Pseudomonas aeruginosa/efectos de los fármacos , Percepción de Quorum/efectos de los fármacos , Factores de Virulencia/antagonistas & inhibidores , Pruebas de Sensibilidad Microbiana , Simulación de Dinámica Molecular , Pseudomonas aeruginosa/fisiología , Metabolismo Secundario

RESUMEN

Staphylococcus aureus, a major cause of nosocomial and community associated infections, is becoming increasingly resistant to antibiotics. S. aureus infections are frequently biofilm-associated, and a diverse spectrum of virulence factors are determinant in the pathogenicity, which can be an interesting target resulting in less selective pressure for bacterial resistance. This study reports anti-virulence activity against S. aureus, including a MRSA clinical isolate, of naphto-γ-pyrones from Aspergillus welwitschiae mycelium, and the characterization of chemical constituents by LC-DAD and LC-MS. A remarkable decrease in biofilm formation, hemolysis and coagulation promoted by Staphylococcus aureus, important traits for bacterial pathogenicity, were observed. Furthermore, advantageous association with vancomycin resulted in significant protection against staphylococcal infection in Galleria mellonella model. These findings corroborate the importance of natural products as a source of new therapeutic possibilities, as well reveals potential alternatives for infection control using anti-virulence and drug association approaches.

Asunto(s)

Antibacterianos/farmacología , Aspergillus/química , Policétidos/farmacología , Staphylococcus aureus/efectos de los fármacos , Vancomicina/farmacología , Factores de Virulencia/antagonistas & inhibidores , Animales , Antibacterianos/administración & dosificación , Biopelículas/efectos de los fármacos , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Larva , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Mariposas Nocturnas/microbiología , Policétidos/aislamiento & purificación , Infecciones Estafilocócicas/tratamiento farmacológico , Vancomicina/administración & dosificación

RESUMEN

The zinc metallopeptidase Pseudomonas elastase (LasB) is a virulence factor of Pseudomonas aeruginosa (P. aeruginosa), a pathogenic bacterium that can cause nosocomial infections. The present study relates the structural analysis of 118 N-alpha-mercaptoacetyl dipeptides (NAMdPs) as LasB inhibitors. Field-based 3D-QSAR and molecular docking methods were employed to describe the essential interactions between NAMdPs and LasB binding sites, and the chemical features that determine their differential activities. We report a predictive 3D-QSAR model that was developed according to the internal and external validation tests. The best model, including steric, electrostatic, hydrogen bond donor, hydrogen bond acceptor, and hydrophobic fields, was found to depict a three-dimensional map with the local positive and negative effects of these chemotypes on the LasB inhibitory activities. Furthermore, molecular docking experiments yielded bioactive conformations of NAMdPs inside the LasB binding site. The series of NAMdPs adopted a similar orientation with respect to phosphoramidon within the LasB binding site (crystallographic reference), where the backbone atoms of NAMdPs are hydrogen-bonded to the LasB residues N112, A113, and R198, similarly to phosphoramidon. Our study also included a deep description of the residues involved in the protein-ligand interaction patterns for the whole set of NAMdPs, through the use of interaction fingerprints (IFPs).

Asunto(s)

Proteínas Bacterianas , Dipéptidos/química , Metaloendopeptidasas , Simulación del Acoplamiento Molecular , Inhibidores de Proteasas/química , Pseudomonas aeruginosa/enzimología , Factores de Virulencia , Proteínas Bacterianas/antagonistas & inhibidores , Proteínas Bacterianas/química , Metaloendopeptidasas/antagonistas & inhibidores , Metaloendopeptidasas/química , Pseudomonas aeruginosa/patogenicidad , Relación Estructura-Actividad Cuantitativa , Factores de Virulencia/antagonistas & inhibidores , Factores de Virulencia/química

RESUMEN

BACKGROUND: Trichophyton rubrum is the main etiological agent of skin and nail infections worldwide. Because of its keratinolytic activity and anthropophilic nature, infection models based on the addition of protein substrates have been employed to assess transcriptional profiles and to elucidate aspects related to host-pathogen interactions. Chalcones are widespread compounds with pronounced activity against dermatophytes. The toxicity of trans-chalcone towards T. rubrum is not fully understood but seems to rely on diverse cellular targets. Within this context, a better understanding of the mode of action of trans-chalcone may help identify new strategies of antifungal therapy and reveal new chemotherapeutic targets. This work aimed to assess the transcriptional profile of T. rubrum grown on different protein sources (keratin or elastin) to mimic natural infection sites and exposed to trans-chalcone in order to elucidate the mechanisms underlying the antifungal activity of trans-chalcone. RESULTS: Overall, the use of different protein sources caused only slight differences in the transcriptional profile of T. rubrum. The main differences were the modulation of proteases and lipases in gene categories when T. rubrum was grown on keratin and elastin, respectively. In addition, some genes encoding heat shock proteins were up-regulated during the growth of T. rubrum on keratin. The transcriptional profile of T. rubrum exposed to trans-chalcone included four main categories: fatty acid and lipid metabolism, overall stress response, cell wall integrity pathway, and alternative energy metabolism. Consistently, T. rubrum Mapk was strongly activated during the first hours of trans-chalcone exposure. Noteworthy, trans-chalcone inhibited genes involved in keratin degradation. The results also showed effects of trans-chalcone on fatty acid synthesis and metabolic pathways involved in acetyl-CoA supply. CONCLUSION: Our results suggest that the mode of action of trans-chalcone is related to pronounced changes in fungal metabolism, including an imbalance between fatty acid synthesis and degradation that interferes with cell membrane and cell wall integrity. In addition, this compound exerts activity against important virulence factors. Taken together, trans-chalcone acts on targets related to dermatophyte physiology and the infection process.

Asunto(s)

Pared Celular/química , Chalcona/farmacología , Ácidos Grasos/metabolismo , Proteínas Fúngicas/metabolismo , Tiña/metabolismo , Trichophyton/metabolismo , Factores de Virulencia/antagonistas & inhibidores , Antifúngicos/farmacología , Pared Celular/genética , Elastina/metabolismo , Proteínas Fúngicas/genética , Perfilación de la Expresión Génica , Regulación Fúngica de la Expresión Génica , Humanos , Queratinas/metabolismo , Transducción de Señal , Tiña/tratamiento farmacológico , Tiña/microbiología , Trichophyton/efectos de los fármacos , Trichophyton/genética

RESUMEN

Antimicrobial resistance constitutes one of the major challenges facing humanity in the Twenty-First century. The spread of resistant pathogens has been such that the possibility of returning to a pre-antibiotic era is real. In this scenario, innovative therapeutic strategies must be employed to restrict resistance. Among the innovative proposed strategies, anti-virulence therapy has been envisioned as a promising alternative for effective control of the emergence and spread of resistant pathogens. This review presents some of the anti-virulence strategies that are currently being developed, it will cover strategies focused on quench pathogen quorum sensing (QS) systems, disassemble of bacterial functional membrane microdomains (FMMs), disruption of biofilm formation and bacterial toxin neutralization.

Asunto(s)

Antibacterianos/farmacología , Bacterias/efectos de los fármacos , Biopelículas/efectos de los fármacos , Descubrimiento de Drogas/tendencias , Microdominios de Membrana/efectos de los fármacos , Percepción de Quorum/efectos de los fármacos , Virulencia/efectos de los fármacos , Antibacterianos/aislamiento & purificación , Bacterias/patogenicidad , Factores de Virulencia/antagonistas & inhibidores

RESUMEN

Cryptococcosis is an opportunistic or primary fungal infection considered to be the most prevalent fatal fungal disease worldwide. Owing to the limited number of available drugs, it is necessary to search for novel antifungal compounds. In the present work, we assessed the antifungal efficacy of three thiazole derivatives (1, 2, and 3). We conducted in vitro and in vivo assays to investigate their effects on important virulence factors, such as capsule and biofilm formation. In addition, the phagocytosis index of murine macrophages exposed to compounds 1, 2, and 3 and the in vivo efficacy of 1, 2, and 3 in Galleria mellonella infected with Cryptococcus spp. were evaluated. All compounds exhibited antifungal activity against biofilms and demonstrated a reduction in biofilm metabolic activity by 43-50% for C. gattii and 26-42% for C. neoformans. Thiazole compounds promoted significant changes in the capsule thickness of C. gattii compared to that of C. neoformans. Further examination of these compounds suggests that they can improve the phagocytosis process of peritoneal murine macrophages in vitro, causing an increase in the phagocytosis rate. Survival percentage was examined in the invertebrate model Galleria mellonella larvae, and only compound 3 could increase the survival at doses of 5 mg/kg after infection with C. gattii (P = .0001) and C. neoformans (P = .0007), similar to fluconazole at 10 mg/kg. The results demonstrated that thiazole compounds, mainly compound 3, have potential to be used for future studies in the search for new therapeutics for cryptococcosis.

Asunto(s)

Antifúngicos/farmacología , Biopelículas/efectos de los fármacos , Criptococosis/microbiología , Cryptococcus/efectos de los fármacos , Cryptococcus/patogenicidad , Tiazoles/farmacología , Factores de Virulencia/antagonistas & inhibidores , Animales , Antifúngicos/química , Biopelículas/crecimiento & desarrollo , Células Cultivadas , Criptococosis/inmunología , Modelos Animales de Enfermedad , Polisacáridos Fúngicos/biosíntesis , Larva/microbiología , Larva/fisiología , Macrófagos/efectos de los fármacos , Macrófagos/microbiología , Ratones , Estructura Molecular , Mariposas Nocturnas , Fagocitosis/efectos de los fármacos , Análisis de Supervivencia , Tiazoles/química

RESUMEN

Staphylococcus aureus is an opportunistic pathogen related to a variety of life-threatening infections but for which antimicrobial resistance is liming the treatment options. We report here that myricetin, but not its glycosylated form, can remarkably decrease the production of several S. aureus virulence factors, including adhesion, biofilm formation, hemolysis and staphyloxanthin production, without interfering with growth. Myricetin affects both surface proteins and secreted proteins which indicate that its action is unrelated to inhibition of the agr quorum sensing system. Analysis of virulence related gene expression and computational simulations of pivotal proteins involved in pathogenesis demonstrate that myricetin downregulates the saeR global regulator and interacts with sortase A and α-hemolysin. Furthermore, Myr confers a significant degree of protection against staphylococcal infection in the Galleria mellonella model. The present findings reveal the potential of Myr as an alternative multi-target antivirulence candidate to control S. aureus pathogenicity.

Asunto(s)

Flavonoides/farmacología , Mariposas Nocturnas/efectos de los fármacos , Mariposas Nocturnas/microbiología , Sustancias Protectoras/farmacología , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/fisiología , Factores de Virulencia/antagonistas & inhibidores , Animales , Adhesión Bacteriana/efectos de los fármacos , Biopelículas/efectos de los fármacos , Eritrocitos/efectos de los fármacos , Eritrocitos/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Flavonoides/química , Perfilación de la Expresión Génica , Hemólisis/efectos de los fármacos , Modelos Moleculares , Unión Proteica , Conformación Proteica , Relación Estructura-Actividad , Xantófilas/biosíntesis , Xantófilas/química

RESUMEN

Decreased antimicrobial efficiency has become a global public health issue. The paucity of new antibacterial drugs is evident, and the arsenal against infectious diseases needs to be improved urgently. The selection of plants as a source of prototype compounds is appropriate, since plant species naturally produce a wide range of secondary metabolites that act as a chemical line of defense against microorganisms in the environment. Although traditional approaches to combat microbial infections remain effective, targeting microbial virulence rather than survival seems to be an exciting strategy, since the modulation of virulence factors might lead to a milder evolutionary pressure for the development of resistance. Additionally, anti-infective chemotherapies may be successfully achieved by combining antivirulence and conventional antimicrobials, extending the lifespan of these drugs. This review presents an updated discussion of natural compounds isolated from plants with chemically characterized structures and activity against the major bacterial virulence factors: quorum sensing, bacterial biofilms, bacterial motility, bacterial toxins, bacterial pigments, bacterial enzymes, and bacterial surfactants. Moreover, a critical analysis of the most promising virulence factors is presented, highlighting their potential as targets to attenuate bacterial virulence. The ongoing progress in the field of antivirulence therapy may therefore help to translate this promising concept into real intervention strategies in clinical areas.

Asunto(s)

Antibacterianos/farmacología , Bacterias/efectos de los fármacos , Productos Biológicos/farmacología , Factores de Virulencia/antagonistas & inhibidores , Bacterias/metabolismo , Fenómenos Fisiológicos Bacterianos , Proteínas Bacterianas/antagonistas & inhibidores , Toxinas Bacterianas/antagonistas & inhibidores , Biopelículas/efectos de los fármacos , Productos Biológicos/aislamiento & purificación , Inhibidores Enzimáticos/farmacología , Pigmentos Biológicos/antagonistas & inhibidores , Plantas/química , Percepción de Quorum

RESUMEN

INTRODUCTION: This is the first study to examine the in vitro susceptibility and the expression of virulence factors in Candida species in the presence of Pimenta pseudocaryophyllus (Gomes) L.R. Landrum (Myrtaceae), a Brazilian plant known as paucravo. Additionally, the mechanisms of action of the crude ethanol extract and the ethyl acetate and aqueous fractions of this plant were investigated. METHODS: The in vitro susceptibility of Candida was tested using the broth microdilution method, whereas an XTT reduction assay was used for biofilms. Adherence was determined by counting the number of yeast cells that adhered to 100 oral epithelial cells, and hyphal formation was verified in the hyphal induction medium M199. Flow cytometry with propidium iodide and FUN-1 was performed to assess the mechanism of action. RESULTS: The results revealed that the crude ethanol extract and the ethyl acetate and aqueous fractions of P. pseudocaryophyllus inhibited the growth of Candida isolates at a minimal inhibitory concentration (MIC) ranging from 64 to 256 µg/mL, whereas the 50% sessile minimal inhibitory concentration (SMIC50) ranged from 512 to >1,024 µg/mL. Adherence and hyphal formation were significantly reduced in the presence of the crude ethanol extract and both fractions. Although cell membrane injury was detected, the predominant mechanism of action appeared to be the alteration of yeast metabolism, as demonstrated by flow cytometry. CONCLUSIONS: Our results indicated that antifungal activity reduced the expression of virulence factors in yeast via the alteration of yeast metabolism, suggesting that the crude extract of P. pseudocaryophyllus and its fractions may contain novel antifungal agents.

Asunto(s)

Antifúngicos/farmacología , Candida/efectos de los fármacos , Pimenta/química , Extractos Vegetales/farmacología , Factores de Virulencia/antagonistas & inhibidores , Antifúngicos/aislamiento & purificación , Biopelículas/efectos de los fármacos , Candida/patogenicidad , Citometría de Flujo , Humanos , Dosificación Letal Mediana , Pruebas de Sensibilidad Microbiana

RESUMEN

Introduction This is the first study to examine the in vitro susceptibility and the expression of virulence factors in Candida species in the presence of Pimenta pseudocaryophyllus (Gomes) L.R. Landrum (Myrtaceae), a Brazilian plant known as paucravo. Additionally, the mechanisms of action of the crude ethanol extract and the ethyl acetate and aqueous fractions of this plant were investigated. Methods The in vitro susceptibility of Candida was tested using the broth microdilution method, whereas an XTT reduction assay was used for biofilms. Adherence was determined by counting the number of yeast cells that adhered to 100 oral epithelial cells, and hyphal formation was verified in the hyphal induction medium M199. Flow cytometry with propidium iodide and FUN-1 was performed to assess the mechanism of action. Results The results revealed that the crude ethanol extract and the ethyl acetate and aqueous fractions of P. pseudocaryophyllus inhibited the growth of Candida isolates at a minimal inhibitory concentration (MIC) ranging from 64 to 256µg/mL, whereas the 50% sessile minimal inhibitory concentration (SMIC50) ranged from 512 to >1,024µg/mL. Adherence and hyphal formation were significantly reduced in the presence of the crude ethanol extract and both fractions. Although cell membrane injury was detected, the predominant mechanism of action appeared to be the alteration of yeast metabolism, as demonstrated by flow cytometry. Conclusions Our results indicated that antifungal activity reduced the expression of virulence factors in yeast via the alteration of yeast metabolism, suggesting that the crude extract of P. pseudocaryophyllus and its fractions may contain novel antifungal agents. .

Asunto(s)

Humanos , Antifúngicos/farmacología , Candida/efectos de los fármacos , Pimenta/química , Extractos Vegetales/farmacología , Factores de Virulencia/antagonistas & inhibidores , Antifúngicos/aislamiento & purificación , Biopelículas/efectos de los fármacos , Candida/patogenicidad , Citometría de Flujo , Pruebas de Sensibilidad Microbiana

RESUMEN

BACKGROUND AND AIMS: Quorum sensing (QS) is a process of bacterial cell-cell communication that controls a large number of systems affecting pathogenicity. Interrupting this communication system can provide nonvirulent pathogenic bacteria. The aim of this study was to evaluate the anti-quorum sensing (anti-QS) potential of an anacardic acids mixture isolated from Amphipterygium adstringens, a medicinal plant known as "cuachalalate", to prevent the onset of bacterial infections as an alternate to antibiotics. METHODS: Initially we investigated the anti-QS activity of A. adstringens hexane extract (HE) by the inhibition of violacein production in Chromobacterium violaceum. From the active HE, an anacardic acid mixture (AAM) was obtained. The anti-quorum sensing activity of AAM was investigated by the rhamnolipid and pyocyanin production constraint as well as decrease of elastase activity, all being quorum sensing-controlled virulence factors expressed in the pathogenic bacteria Pseudomonas aeruginosa. RESULTS: HE induced a 91.6% of inhibition of the violecin production at 55 µg/mL concentration, whereas AAM showed 94% of inhibition at 166 µg/mL. In both cases, inhibition of violacein production did not affect the viability of the bacterium. AAM inhibited pyocyanin (86% at 200 µg/mL) and rhamnolipid (91% at 500 µg/mL) production in a dose/response form and decrease the elastase (75% at 500 µg/mL) activity in P. aeruginosa without affecting its development. CONCLUSIONS: Because an anacardic acids mixture isolated from A. adstringens demonstrated anti-QS, it could be further exploited for novel molecules to treat the emerging infections of antibiotic-resistant bacterial pathogens.

Asunto(s)

Ácidos Anacárdicos/farmacología , Chromobacterium/efectos de los fármacos , Pseudomonas aeruginosa/efectos de los fármacos , Percepción de Quorum/efectos de los fármacos , Factores de Virulencia/antagonistas & inhibidores , Anacardiaceae/química , Ácidos Anacárdicos/aislamiento & purificación , Animales , Antibacterianos/farmacología , Chromobacterium/crecimiento & desarrollo , Chromobacterium/patogenicidad , Farmacorresistencia Bacteriana , Glucolípidos/antagonistas & inhibidores , Glucolípidos/metabolismo , Indoles/antagonistas & inhibidores , Indoles/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Pseudomonas aeruginosa/crecimiento & desarrollo , Pseudomonas aeruginosa/patogenicidad , Piocianina/antagonistas & inhibidores , Piocianina/metabolismo , Factores de Virulencia/metabolismo

RESUMEN

The objective of this study was to evaluate whether Candida albicans exhibits altered pathogenicity characteristics following sublethal antimicrobial photodynamic inactivation (APDI) and if such alterations are maintained in the daughter cells. C. albicans was exposed to sublethal APDI by using methylene blue (MB) as a photosensitizer (0.05 mM) combined with a GaAlAs diode laser (λ 660 nm, 75 mW/cm(2), 9 to 27 J/cm(2)). In vitro, we evaluated APDI effects on C. albicans growth, germ tube formation, sensitivity to oxidative and osmotic stress, cell wall integrity, and fluconazole susceptibility. In vivo, we evaluated C. albicans pathogenicity with a mouse model of systemic infection. Animal survival was evaluated daily. Sublethal MB-mediated APDI reduced the growth rate and the ability of C. albicans to form germ tubes compared to untreated cells (P < 0.05). Survival of mice systemically infected with C. albicans pretreated with APDI was significantly increased compared to mice infected with untreated yeast (P < 0.05). APDI increased C. albicans sensitivity to sodium dodecyl sulfate, caffeine, and hydrogen peroxide. The MIC for fluconazole for C. albicans was also reduced following sublethal MB-mediated APDI. However, none of those pathogenic parameters was altered in daughter cells of C. albicans submitted to APDI. These data suggest that APDI may inhibit virulence factors and reduce in vivo pathogenicity of C. albicans. The absence of alterations in daughter cells indicates that APDI effects are transitory. The MIC reduction for fluconazole following APDI suggests that this antifungal could be combined with APDI to treat C. albicans infections.

Asunto(s)

Candida albicans/efectos de los fármacos , Candidiasis/tratamiento farmacológico , Azul de Metileno/farmacología , Fármacos Fotosensibilizantes/farmacología , Factores de Virulencia/antagonistas & inhibidores , Animales , Antifúngicos/farmacología , Candida albicans/crecimiento & desarrollo , Candida albicans/patogenicidad , Candida albicans/efectos de la radiación , Candidiasis/microbiología , Candidiasis/mortalidad , Pared Celular/efectos de los fármacos , Pared Celular/efectos de la radiación , Femenino , Fluconazol/farmacología , Patrón de Herencia , Láseres de Semiconductores , Luz , Masculino , Ratones , Ratones Endogámicos BALB C , Presión Osmótica , Estrés Oxidativo , Análisis de Supervivencia , Factores de Virulencia/metabolismo

RESUMEN

Trans-sialidase (TS), a virulence factor from Trypanosoma cruzi, is an enzyme playing key roles in the biology of this protozoan parasite. Absent from the mammalian host, it constitutes a potential target for the development of novel chemotherapeutic drugs, an urgent need to combat Chagas' disease. TS is involved in host cell invasion and parasite survival in the bloodstream. However, TS is also actively shed by the parasite to the bloodstream, inducing systemic effects readily detected during the acute phase of the disease, in particular, hematological alterations and triggering of immune cells apoptosis, until specific neutralizing antibodies are elicited. These antibodies constitute the only known submicromolar inhibitor of TS's catalytic activity. We now report the identification and detailed characterization of a neutralizing mouse monoclonal antibody (mAb 13G9), recognizing T. cruzi TS with high specificity and subnanomolar affinity. This mAb displays undetectable association with the T. cruzi superfamily of TS-like proteins or yet with the TS-related enzymes from Trypanosoma brucei or Trypanosoma rangeli. In immunofluorescence assays, mAb 13G9 labeled 100% of the parasites from the infective trypomastigote stage. This mAb also reduces parasite invasion of cultured cells and strongly inhibits parasite surface sialylation. The crystal structure of the mAb 13G9 antigen-binding fragment in complex with the globular region of T. cruzi TS was determined, revealing detailed molecular insights of the inhibition mechanism. Not occluding the enzyme's catalytic site, the antibody performs a subtle action by inhibiting the movement of an assisting tyrosine (Y119), whose mobility is known to play a key role in the trans-glycosidase mechanism. As an example of enzymatic inhibition involving non-catalytic residues that occupy sites distal from the substrate-binding pocket, this first near atomic characterization of a high affinity inhibitory molecule for TS provides a rational framework for novel strategies in the design of chemotherapeutic compounds.

Asunto(s)

Anticuerpos Monoclonales de Origen Murino/química , Anticuerpos Neutralizantes/química , Glicoproteínas/química , Neuraminidasa/química , Trypanosoma cruzi/enzimología , Factores de Virulencia/química , Animales , Anticuerpos Monoclonales de Origen Murino/inmunología , Anticuerpos Monoclonales de Origen Murino/uso terapéutico , Anticuerpos Neutralizantes/inmunología , Anticuerpos Neutralizantes/uso terapéutico , Sitios de Unión , Enfermedad de Chagas/tratamiento farmacológico , Enfermedad de Chagas/enzimología , Glicoproteínas/antagonistas & inhibidores , Glicoproteínas/inmunología , Ratones , Neuraminidasa/antagonistas & inhibidores , Neuraminidasa/inmunología , Estructura Cuaternaria de Proteína , Trypanosoma cruzi/inmunología , Trypanosoma cruzi/patogenicidad , Factores de Virulencia/antagonistas & inhibidores , Factores de Virulencia/inmunología

RESUMEN

As well as the host, opportunist Candida spp. enface all sorts of exogenous chemicals, so-called xenobiotics. It is plausible that xenobiotics exert some effects on such microorganisms; among them, the modulation of virulence attributes.

Asunto(s)

Antifúngicos/farmacología , Candida/efectos de los fármacos , Factores de Virulencia/antagonistas & inhibidores , Xenobióticos/farmacología , Humanos , Virulencia

RESUMEN

BACKGROUND: Paracoccidioides brasiliensis is a human thermal dimorphic pathogenic fungus. Survival of P. brasiliensis inside the host depends on the adaptation of this fungal pathogen to different conditions, namely oxidative stress imposed by immune cells. AIMS AND METHODOLOGY: In this study, we evaluated the role of alternative oxidase (AOX), an enzyme involved in the intracellular redox balancing, during host-P. brasiliensis interaction. We generated a mitotically stable P. brasiliensis AOX (PbAOX) antisense RNA (aRNA) strain with a 70% reduction in gene expression. We evaluated the relevance of PbAOX during interaction of conidia and yeast cells with IFN-γ activated alveolar macrophages and in a mouse model of infection. Additionally, we determined the fungal cell's viability and PbAOX in the presence of H2O2. RESULTS: Interaction with IFN-γ activated alveolar macrophages induced higher levels of PbAOX gene expression in PbWt conidia than PbWt yeast cells. PbAOX-aRNA conidia and yeast cells had decreased viability after interaction with macrophages. Moreover, in a mouse model of infection, we showed that absence of wild-type levels of PbAOX in P. brasiliensis results in a reduced fungal burden in lungs at weeks 8 and 24 post-challenge and an increased survival rate. In the presence of H2O2, we observed that PbWt yeast cells increased PbAOX expression and presented a higher viability in comparison with PbAOX-aRNA yeast cells. CONCLUSIONS: These data further support the hypothesis that PbAOX is important in the fungal defense against oxidative stress imposed by immune cells and is relevant in the virulence of P. brasiliensis.

Asunto(s)

Viabilidad Microbiana , Proteínas Mitocondriales/metabolismo , Oxidorreductasas/metabolismo , Paracoccidioides/enzimología , Paracoccidioides/patogenicidad , Proteínas de Plantas/metabolismo , Factores de Virulencia/metabolismo , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Técnicas de Silenciamiento del Gen , Peróxido de Hidrógeno/toxicidad , Pulmón/microbiología , Macrófagos/inmunología , Macrófagos/microbiología , Masculino , Ratones , Ratones Endogámicos BALB C , Proteínas Mitocondriales/antagonistas & inhibidores , Proteínas Mitocondriales/genética , Oxidantes/toxicidad , Oxidorreductasas/antagonistas & inhibidores , Oxidorreductasas/genética , Paracoccidioides/efectos de los fármacos , Paracoccidioides/inmunología , Paracoccidioidomicosis/microbiología , Paracoccidioidomicosis/patología , Proteínas de Plantas/antagonistas & inhibidores , Proteínas de Plantas/genética , ARN sin Sentido/genética , ARN sin Sentido/metabolismo , Análisis de Supervivencia , Virulencia , Factores de Virulencia/antagonistas & inhibidores , Factores de Virulencia/genética

RESUMEN

Candida yeasts are opportunistic pathogens responsible for infections in immunocompromised individuals. Among the virulence factors present in these yeasts we can mention the ability to adhere to host cells, exoenzyme production and germ tube formation. Several compounds, such as antifungal agents, plants extracts, protein inhibitors and surfactants, have been tested regarding their capacity in inhibit Candida spp. virulence factors. Among these compounds, a significant lower number of works are focused on the inhibition action caused by different types of surfactant. The present work aimed to evaluate the effect generated by the surfactants cetyltrimethylammonium chloride (CTAC), sodium dodecyl sulfate (SDS), N-hexadecyl-N-N'-dimethyl-3-ammonio-1-propane-sulfonate (HPS) and octylphenoxypolyethoxyethanol (Triton X-100) on the viability, adhesion ability and exoenzyme production by Candida species. CTAC and HPS were capable to inhibit Candida spp. growth at very low concentrations. All surfactants demonstrated to be capable to inhibit the adhesion of Candida species to buccal epithelial cells (BEC) and the proteinase production. On the other hand, the phospholipase production remained unaltered after the treatment with these compounds. The present data denote that cationic and zwitterionic surfactants are interesting prototypes of inhibitory agents against Candida spp., which is probably associated with the cationic punctual charge of both surfactants. The results are discussed in details in agreement with recent reports from literature.

Asunto(s)

Antifúngicos/farmacología , Candida/efectos de los fármacos , Candida/patogenicidad , Tensoactivos/farmacología , Factores de Virulencia/antagonistas & inhibidores , Adulto , Candida/crecimiento & desarrollo , Candida/metabolismo , Adhesión Celular/efectos de los fármacos , Células Cultivadas , Endopeptidasas/metabolismo , Células Epiteliales/microbiología , Humanos , Viabilidad Microbiana/efectos de los fármacos , Fosfolipasas/antagonistas & inhibidores

RESUMEN

Toxigenic strains of Clostridium difficile were co-cultured with different strains of bifidobacteria and lactobacilli. Spent culture supernatants were tested for biological activity on cultured Vero cells. Co-culture of C. difficile with some potentially probiotic strains lead to a reduction of the biological activity of spent culture supernatants. The observed effects cannot be ascribed either to secreted factors from the probiotic strains or to toxin adsorption by bacterial cells. Immunological assays showed that there was significant diminution of both clostridial toxins (TcdA and TcdB) in spent culture supernatants of co-cultures as compared with pure clostridial cultures. Even though co-cultured clostridial cells showed a slight increase of intracellular toxins, this increase did not completely explains the reduction of toxin concentration in culture supernatants. The evidence suggests that the antagonism could be due to the diminution of the synthesis and/or secretion of both clostridial toxins. Our findings provide new insights into the possible mechanisms involved in the protective effect of probiotics in the context of C. difficile infection.

Asunto(s)

Antibiosis , Clostridioides difficile/metabolismo , Probióticos/farmacología , Factores de Virulencia/antagonistas & inhibidores , Proteínas Bacterianas/antagonistas & inhibidores , Toxinas Bacterianas/antagonistas & inhibidores , Clostridioides difficile/crecimiento & desarrollo , Técnicas de Cocultivo , Enterotoxinas/antagonistas & inhibidores