9 research outputs found
Staphylococcus aureus RnpA Inhibitors: Computational-Guided Design, Synthesis and Initial Biological Evaluation
No full textAntibiotic resistance is spreading worldwide and it has become one of the most important issues in modern medicine. In this context, the bacterial RNA degradation and processing machinery are essential processes for bacterial viability that may be exploited for antimicrobial therapy. In Staphylococcus aureus, RnpA has been hypothesized to be one of the main players in these mechanisms. S. aureus RnpA is able to modulate mRNA degradation and complex with a ribozyme (rnpB), facilitating ptRNA maturation. Corresponding small molecule screening campaigns have recently identified a few classes of RnpA inhibitors, and their structure activity relationship (SAR) has only been partially explored. Accordingly, in the present work, using computational modeling of S. aureus RnpA we identified putative crucial interactions of known RnpA inhibitors, and we used this information to design, synthesize, and biologically assess new potential RnpA inhibitors. The present results may be beneficial for the overall knowledge about RnpA inhibitors belonging to both RNPA2000-like thiosemicarbazides and JC-like piperidine carboxamides molecular classes. We evaluated the importance of the different key moieties, such as the dichlorophenyl and the piperidine of JC2, and the semithiocarbazide, the furan, and the i-propylphenyl ring of RNPA2000. Our efforts could provide a foundation for further computational-guided investigations
Comparative genomic analysis of European and Middle Eastern community-associated methicillin-resistant Staphylococcus aureus (CC80:ST80-IV) isolates by high-density microarray
No full textAbstractInfections as a result of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) are an issue of increasing global healthcare concern. In Europe, this principally involves strains of multi-locus sequence type clonal complex 80 sequence type 80 with methicillin resistance in a staphylococcal chromosomal cassette (SCCmec) type IV arrangement (CC80:ST80-IV). As with other CA-MRSA strains, CC80:ST80-IV isolates tend to appear uniform when analysed by common molecular typing methods (e.g. pulsed field gel electrophoresis, multi-locus sequence type, SCCmec). To explore whether DNA sequence-based differences exist, we compared the genetic composition of six CC80:ST80-IV isolates of diverse chronological and geographic origin (i.e. Denmark and the Middle East) using an Affymetrix high-density microarray that was previously used to analyse CA-MRSA USA300 isolates. The results revealed a high degree of homology despite the diversity in isolation date and origin, with isolate differences primarily in conserved hypothetical open reading frames and intergenic sequences, but also including regions of known function. This included the confirmed loss of SCCmec recombinase genes in two Danish isolates representing potentially new SCCmec types. Microarray analysis grouped the six isolates into three relatedness pairs, also identified by pulsed field gel electrophoresis, which were consistent with both the clinical and molecular data
Drug-Eluting Cements for Hard Tissue Repair: A Comparative Study using Vancomycin and RNPA1000 to Inhibit Growth of Staphylococcus Aureus
No full textBone cement used in orthopaedic applications can become colonized with bacterial biofilms, resulting in severe medical complications. Consequently, bone cements are often loaded with antibiotics in an effort to prevent bacterial colonization. However, current formulations may not release antibiotics into the environment at sufficient and sustained concentrations required to impede bacterial growth or may be incompatible with antibiotics that are effective against the colonizing organism. Thus, new cement formulation options are needed. This report describes the performance of a novel SiO2-TiO2-ZnO-CaO-SrO- based glass polyalkenoate cement as a carrier of antimicrobials active against Staphylococcus aureus, the predominant cause of orthopaedic biofilm-associated infections. The antibiotic vancomycin and a novel Staphylococcus aureus RnpA inhibitor under pre-clinical development, RNPA1000, were included in these studies. Rheological testing characterized the workability of the glass polyalkenoate cement over a range of powder-to-liquid ratios and polyacrylic acid concentrations and revealed that the most suitable powder-to-liquid ratio was 2/1.25 with 40 wt& polyacrylic acid. Loading glass polyalkenoate cement with either 20-30& RNPA1000 or vancomycin prevented bacterial growth. However, longer incubations allowed for Staphylococcus aureus colonies to form near the vancomycin-infused cement, indicating that vancomycin may not be suitable for long-term biofilm inhibition in comparison to RNPA1000. Scanning electron microscopy and energy-dispersive X-ray analyses confirmed successful incorporation RNPA1000 into the cement matrix and were indicative of its slow release. These studies establish a drug-eluting formulation of glass polyalkenoate cement with great potential in orthopaedic implants that incorporates known antibiotics as well as RNPA1000 to prevent growth of the dangerous pathogen Staphylococcus aureus. © The Author(s) 2013 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav
Role of sigmaB in the expression of Staphylococcus aureus cell wall adhesins ClfA and FnbA and contribution to infectivity in a rat model of experimental endocarditis
No full textIsogenic Staphylococcus aureus strains with different capacities to produce sigma(B) activity were analyzed for their ability to attach to fibrinogen- or fibronectin-coated surfaces or platelet-fibrin clots and to cause endocarditis in rats. In comparison to the sigma(B)-deficient strain, BB255, which harbors an rsbU mutation, both rsbU-complemented and sigma(B)-overproducing derivatives exhibited at least five times greater attachment to fibrinogen- and fibronectin-coated surfaces and showed increased adherence to platelet-fibrin clots. No differences in adherence were seen between BB255 and a DeltarsbUVWsigB isogen. Northern blotting analyses revealed that transcription of clfA, encoding fibrinogen-binding protein clumping factor A, and fnbA, encoding fibronectin-binding protein A, were positively influenced by sigma(B). Sigma(B) overproduction resulted in a statistically significant increase in positive spleen cultures and enhanced bacterial densities in both the aortic vegetations and spleens at 16 h postinoculation. In contrast, at 72 h postinoculation, tissues infected with the sigma(B) overproducer had lower bacterial densities than did those infected with BB255. These results suggest that although sigma(B) appears to increase the adhesion of S. aureus to various host cell-matrix proteins in vitro, it has limited effect on pathogenesis in the rat endocarditis model. Sigma(B) appears to have a transient enhancing effect on bacterial density in the early stages of infection that is lost during progression
Difference in virulence between Staphylococcus aureus isolates causing gangrenous mastitis versus subclinical mastitis in a dairy sheep flock
No full textStaphylococcus aureus mastitis in dairy sheep ranges from subclinical mastitis to lethal gangrenous mastitis. Neither the S. aureus virulence factors nor the host-factors or the epidemiological events contributing to the different outcomes are known. In a field study in a dairy sheep farm over 21 months, 16 natural isolates of S. aureus were collected from six subclinical mastitis cases, one lethal gangrenous mastitis case, nasal carriage from eight ewes and one isolate from ambient air in the milking room. A genomic comparison of two strains, one responsible for subclinical mastitis and one for lethal gangrenous mastitis, was performed using multi-strain DNA microarrays. Multiple typing techniques (pulsed-field-gel-electrophoresis, multiple-locus variable-number, single-nucleotide polymorphisms, randomly amplified polymorphic DNA, spa typing and sas typing) were used to characterise the remaining isolates and to follow the persistence of the gangrenous isolate in ewes' nares. Our results showed that the two strains were genetically closely related and they shared 3 615 identical predicted open reading frames. However, the gangrenous mastitis isolate carried variant versions of several genes (sdrD, clfA-B, sasA, sasB, sasD, sasI and splE) and was missing fibrinogen binding protein B (fnbB) and a prophage. The typing results showed that this gangrenous strain emerged after the initial subclinical mastitis screening, but then persisted in the flock in the nares of four ewes. Although we cannot dismiss the role of host susceptibility in the clinical events in this flock, our data support the hypothesis that S. aureus populations had evolved in the sheep flock and that S. aureus genetic variations could have contributed to enhanced virulence
Impact of target site distribution for Type I restriction enzymes on the evolution of methicillin-resistant Staphylococcus aureus (MRSA) populations.
No full textA limited number of Methicillin-resistant Staphylococcus aureus (MRSA) clones are responsible for MRSA infections worldwide, and those of different lineages carry unique Type I restriction-modification (RM) variants. We have identified the specific DNA sequence targets for the dominant MRSA lineages CC1, CC5, CC8 and ST239. We experimentally demonstrate that this RM system is sufficient to block horizontal gene transfer between clinically important MRSA, confirming the bioinformatic evidence that each lineage is evolving independently. Target sites are distributed randomly in S. aureus genomes, except in a set of large conjugative plasmids encoding resistance genes that show evidence of spreading between two successful MRSA lineages. This analysis of the identification and distribution of target sites explains evolutionary patterns in a pathogenic bacterium. We show that a lack of specific target sites enables plasmids to evade the Type I RM system thereby contributing to the evolution of increasingly resistant community and hospital MRSA
Rapeseed oil spray development of diesel idi spray nozzle under air movement influence
No full textRapeseed oil (RO) spray has very slow atomization due to its high viscosity nature. Although high injection pressure, high ambient temperature and combination of nozzle can promote faster atomization of rapeseed oil spray, another factor that was not discussed is the effect air movement that could positively influence RO spray development. To study the effect of air movement, in particular to generate the swirl (air movement inside the chamber), a swirler was used. Images were captured using a nano-spark shadowgraph photography technique and also high speed video imaging. Macrostructures of diesel sprays such as spray tip penetration length, spray shape, spraycone angle were obtained. Microstructures, such as droplet distribution and size were also studied. Result shows that IDI nozzle rapeseed oil spray has a narrow spray cone angle. The average droplet size is around 20~25µm. The large size of rapeseed oil droplet require assistant to improve atomization and results shows rapid air movement in chamber successfully improve atomizatio
Rol del T6SS en Competencia Bacteriana de Xanthomonas phaseoli pv. manihotis Ante Bacterias Epífitas de Yuca (Manihot esculenta)
No full textMediante el Sistema de secreción tipo VI (T6SS), las bacterias gramnegativas pueden translocar proteínas efectoras que juegan un papel importante en competencia bacteriana y virulencia hacia hospederos eucariotas. En este estudio, fue posible realizar el aislamiento e identificación taxonómica mediante secuenciación del gen 16S, de seis bacterias epífitas que habitan la filósfera de yuca (Manihot esculenta), una especie vegetal de importancia económica en Sur América y África. Ensayos de competencia bacteriana in vitro sugirieron que el T6SS de la bacteria fitopatógena Xanthomonas phaseoli pv. manihotis (Xpm) era requerido por contacto directo para ejercer competencia bacteriana hacia la bacteria epífita de yuca Rhodococcus sp. Mutantes para los genes VgrG, ClpV y Hcp del T6SS de Xpm fueron incapaces de inhibir el crecimiento de Rhodococcus sp. Sin embargo, no fue posible determinar con certeza que el T6SS de Xpm sea el responsable de la inhibición de Rhodococcus sp., debido a la falta de complementación observada de estos mutantes en Xpm. Finalmente, fue posible predecir a través de análisis bioinformáticos dos proteínas efectoras que serían hipotéticamente traslocadas por el T6SS de Xpm, y el papel que podrían cumplir en la competencia bacteriana. Los resultados obtenidos en este estudio ayudan a comprender de alguna manera, cómo el T6SS podría jugar un papel importante en competencia bacteriana posibilitando que Xpm gane ventaja de nicho al matar a otras bacterias, facilitando finalmente el establecimiento de la enfermedad tizón bacteriano de la yuca (CBB).Through the Type VI Secretion System (T6SS), Gram-negative bacteria can translocate effector proteins that play an important role in bacterial competition and virulence towards eukaryotic hosts. In this study, it was possible to perform the isolation and taxonomic identification through 16S gene sequencing of six epiphytic bacteria that inhabit the cassava (Manihot esculenta) phyllosphere, a plant species of economic importance in South America and Africa. In vitro bacterial competition assays suggested that T6SS from the phytopathogenic bacterium Xanthomonas phaseoli pv. manihotis (Xpm) was required by direct contact to exert bacterial competition towards the cassava epiphytic bacterium Rhodococcus sp. Mutants for the VgrG, ClpV and Hcp genes of the Xpm T6SS were unable to inhibit the growth of Rhodococcus sp. However, it was not possible to determine with certainty that the T6SS of Xpm is responsible for the inhibition of Rhodococcus sp., due to the observed lack of complementation of these mutants in Xpm. Finally, it was possible to predict through bioinformatic analysis two effector proteins that would be hypothetically translocated by the T6SS of Xpm, and the role that they could play in bacterial competition. The results obtained in this study help to understand in some way, how T6SS could play an important role in bacterial competition, enabling Xpm to gain a niche advantage by killing other bacteria, finally facilitating the establishment of cassava bacterial blight disease (CBB).Magíster en Ciencias BiológicasMaestríaFitopatología Molecula
