Heterogeneous vancomycin-intermediate susceptibility in a community-associated methicillin-resistant Staphylococcus aureus epidemic clone, in a case of Infective Endocarditis in Argentina

Background

There has been considerable effort to discover plant-derived antibacterials against methicillin-resistant strains of Staphylococcus aureus (MRSA) which have developed resistance to most existing antibiotics, including the last line of defence, vancomycin. Pentacyclic triterpenoid, a biologically diverse plant-derived natural product, has been reported to show anti-staphylococcal activities. The objective of this study is to evaluate the interaction between three pentacyclic triterpenoid and standard antibiotics (methicillin and vancomycin) against reference strains of Staphylococcus aureus.

Methods and Results

The activity of the standard antibiotics and compounds on reference methicillin-sensitive and resistant strains of S. aureus were determined using the macrodilution broth method. The minimum inhibitory concentration (MIC) of the compounds was compared with that of the standard antibiotics. The interaction between any two antimicrobial agents was estimated by calculating the fractional inhibitory concentration (FIC index) of the combination. The various combinations of antibiotics and compounds reduced the MIC to a range of 0.05 to 50%.

Conclusion

Pentacyclic triterpenoids have shown anti-staphylococcal activities and although individually weaker than common antibiotics produced from bacteria and fungi, synergistically these compounds may use different mechanism of action or pathways to exert their antimicrobial effects, as implicated in the lowered MICs. Therefore, the use of current antibiotics could be maintained in their combination with plant-derived antibacterial agents as a therapeutic option in the treatment of S. aureus infections.     

Establishment and characterization of atypical fibroblasts from human adult liver contributing to hepatocyte cord-like arrangement

We report the phenotypic and functional characterization of fibroblasts established in culture from the non-parenchymal epithelial cell populations of adult human livers. Human liver fibroblasts (hLF) expressed mesenchymal antigens vimentin, alpha-smooth muscle actin, collagen, fibronectin, CD73, CD90, CD105, and CD166 together with non-mesenchymal antigens cytokeratins 8 and 18, glial fibrillary acidic protein, and nestin. Mixed cell lineage-specific protein expression was not associated with stem-like cell properties. Coculturing hepatocytes onto confluent hLF showed that they survived and maintained metabolic activity such as albumin, glycogen, and urea production. Moreover, hepatocytes formed cord-like arrangements resembling those established in vivo. Hepatocyte arrangement depended on cell-to-cell contact and the tissue origin of fibroblasts. Time-lapse video imaging of cocultured cells showed that hepatocyte arrangement was coordinated by the stretching and shortening of underneath hLF. Our data suggest that hLF may represent resident fibroblasts of the adult human liver, which could assume guiding functions for hepatic epithelial cells.     

Genomic data reveal deep genetic structure but no support for current taxonomic designation in a grasshopper species complex

Taxonomy has traditionally relied on morphological and ecological traits to interpret and classify biological diversity. Over the last decade, technological advances and conceptual developments in the field of molecular ecology and systematics have eased the generation of genomic data and changed the paradigm of biodiversity analysis. Here we illustrate how traditional taxonomy has led to species designations that are supported neither by high throughput sequencing data nor by the quantitative integration of genomic information with other sources of evidence. Specifically, we focus on Omocestus antigai and Omocestus navasi, two montane grasshoppers from the Pyrenean region that were originally described based on quantitative phenotypic differences and distinct habitat associations (alpine vs. Mediterranean‐montane habitats). To validate current taxonomic designations, test species boundaries, and understand the factors that have contributed to genetic divergence, we obtained phenotypic (geometric morphometrics) and genome‐wide SNP data (ddRADSeq) from populations covering the entire known distribution of the two taxa. Coalescent‐based phylogenetic reconstructions, integrative Bayesian model‐based species delimitation, and landscape genetic analyses revealed that populations assigned to the two taxa show a spatial distribution of genetic variation that do not match with current taxonomic designations and is incompatible with ecological/environmental speciation. Our results support little phenotypic variation among populations and a marked genetic structure that is mostly explained by geographic distances and limited population connectivity across the abrupt landscapes characterizing the study region. Overall, this study highlights the importance of integrative approaches to identify taxonomic units and elucidate the evolutionary history of species.     

Alteration of Plasma Membrane Organization by an Anticancer Lysophosphatidylcholine Analogue Induces Intracellular Acidification and Internalization of Plasma Membrane Transporters in Yeast

The lysophosphatidylcholine analogue edelfosine is a potent antitumor lipid that targets cellular membranes. The underlying mechanisms leading to cell death remain controversial, although two cellular membranes have emerged as primary targets of edelfosine, the plasma membrane (PM) and the endoplasmic reticulum. In an effort to identify conditions that enhance or prevent the cytotoxic effect of edelfosine, we have conducted genome-wide surveys of edelfosine sensitivity and resistance in Saccharomyces cerevisiae presented in this work and the accompanying paper (Cuesta-Marbán, Á., Botet, J., Czyz, O., Cacharro, L. M., Gajate, C., Hornillos, V., Delgado, J., Zhang, H., Amat-Guerri, F., Acuña, A. U., McMaster, C. R., Revuelta, J. L., Zaremberg, V., and Mollinedo, F. (January 23, 2013) J. Biol. Chem. 288,), respectively. Our results point to maintenance of pH homeostasis as a major player in modulating susceptibility to edelfosine with the PM proton pump Pma1p playing a main role. We demonstrate that edelfosine alters PM organization and induces intracellular acidification. Significantly, we show that edelfosine selectively reduces lateral segregation of PM proteins like Pma1p and nutrient H⁺-symporters inducing their ubiquitination and internalization. The biology associated to the mode of action of edelfosine we have unveiled includes selective modification of lipid raft integrity altering pH homeostasis, which in turn regulates cell growth.     

Development and validation of a method to determine the unbound paclitaxel fraction in human plasma

Paclitaxel is pharmaceutically formulated in a mixture of Cremophor EL and ethanol (1:1, v/v). The unbound fraction of the anticancer drug paclitaxel in plasma is dependent on both plasma protein binding and entrapment in Cremophor EL micelles. We have developed a simple and reproducible method for the quantification of the unbound paclitaxel fraction in human plasma. Human plasma was spiked with [3H]paclitaxel and [14C]glucose (unbound reference) and incubated at 37 degrees C for 30 min. Plasma ultrafiltrate was prepared by a micropartition system (MPS-1) and collected in a sample cup containing 100 microl of plasma to prevent the loss of paclitaxel due to adsorption. The radionuclides were separated after combustion of the biological samples using a sample oxidizer and the radioactivity was determined by liquid scintillation counting. The unbound fraction of paclitaxel was calculated by dividing the ratios of 3H and 14C in plasma ultrafiltrate and in plasma. The method was thoroughly validated using human plasma spiked with pharmacologically relevant concentrations of paclitaxel (10-1000 ng/ml) and Cremophor EL (0.25-2.0%). The method was precise, with a within-day precision ranging from 3.9 to 11.0% and a between-day precision ranging from 5.8 to 13.1%. In patient plasma with low serum albumin values containing 1% of Cremophor EL, the unbound fraction appeared to be significantly higher than that in plasma with normal albumin values. The determination of the unbound fraction of paclitaxel proved to be stable during a 10-week storage at -20 degrees C. Furthermore, the assay was applicable in patient samples. This assay can be used to determine the unbound fraction of paclitaxel in plasma. Moreover, its design should allow the determination of the unbound concentrations of other hydrophobic drugs.     

Combining Computational and Social Effort for Collaborative Problem Solving

Rather than replacing human labor, there is growing evidence that networked computers create opportunities for collaborations of people and algorithms to solve problems beyond either of them. In this study, we demonstrate the conditions under which such synergy can arise. We show that, for a design task, three elements are sufficient: humans apply intuitions to the problem, algorithms automatically determine and report back on the quality of designs, and humans observe and innovate on others’ designs to focus creative and computational effort on good designs. This study suggests how such collaborations should be composed for other domains, as well as how social and computational dynamics mutually influence one another during collaborative problem solving.     

Phenotypic and phylogenetic characterization of native peanut Bradyrhizobium isolates obtained from Córdoba, Argentina

Peanut is an economically important legume nodulated by slow-growing bacteria of the genus Bradyrhizobium. In this study, a collection of native slow-growing peanut rhizobial isolates from Argentina was obtained and characterized. The phenotypical characterization included the determination of the symbiotic properties, whereas the genetic and phylogenetic diversity was assessed through ERIC-PCR and sequencing of the internal transcribed spacer (ITS) region, as well as the dnaK and nodA genes. The results obtained indicated that peanut nodulating bradyrhizobia were phenotypically and genotypically diverse, and included locally adapted variants of B. yuanmingense and B. iriomotense carrying novel nodA alleles.