Dynamics of bacterial growth and distribution within the liver during Salmonella infection

Salmonella enterica causes severe systemic diseases in humans and animals and grows intracellularly within discrete tissue foci that become pathological lesions. Because of its lifestyle Salmonella is a superb model for studying the in vivo dynamics of bacterial distribution. Using multicolour fluorescence microscopy in the mouse typhoid model we have studied the interaction between different bacterial populations in the same host as well as the dynamic evolution of foci of infection in relation to bacterial growth and localization. We showed that the growth of Salmonella in the liver results in the spread of the microorganisms to new foci of infection rather than simply in the expansion of the initial ones. These foci were associated with independently segregating bacterial populations and with low numbers of bacteria in each infected phagocyte. Using fast-growing and slow-growing bacteria we also showed that the increase in the number of infected phagocytes parallels the net rate of bacterial growth of the microorganisms in the tissues. These findings suggest a novel mechanism underlying growth of salmonellae in vivo with important consequences for understanding mechanisms of resistance and immunity.     

Alterations in the proteome of the respiratory tract in response to single and multiple exposures to naphthalene

Protein adduction is considered to be critical to the loss of cellular homeostasis associated with environmental chemicals undergoing metabolic activation. Despite considerable effort, our understanding of the key proteins mediating the pathologic consequences from protein modification by electrophiles is incomplete. This work focused on naphthalene (NA) induced acute injury of respiratory epithelial cells and tolerance which arises after multiple toxicant doses to define the initial cellular proteomic response and later protective actions related to tolerance. Airways and nasal olfactory epithelium from mice exposed to 15 ppm NA either for 4 h (acute) or for 4 h/day × 7 days (tolerant) were used for label‐free protein quantitation by LC/MS/MS. Cytochrome P450 2F2 and secretoglobin 1A1 are decreased dramatically in airways of mice exposed for 4 h, a finding consistent with the fact that CYPs are localized primarily in Clara cells. A number of heat shock proteins and protein disulfide isomerases, which had previously been identified as adduct targets for reactive metabolites from several lung toxicants, were upregulated in airways but not olfactory epithelium of tolerant mice. Protein targets that are upregulated in tolerance may be key players in the pathophysiology associated with reactive metabolite protein adduction. All MS data have been deposited in the ProteomeXchange with identifier PXD000846 ( http://proteomecentral.proteomexchange.org/dataset/PXD000846 ).     

Thrombospondin‐1 is part of a Slug‐independent motility and metastatic program in cutaneous melanoma,in association with VEGFR‐1 and FGF‐2

Differently from most transformed cells, cutaneous melanoma expresses the pleiotropic factor thrombospondin‐1 (TSP‐1). Herein, we show that TSP‐1 (RNA and protein), undetectable in four cultures of melanocytes and a RGP melanoma, was variously present in 13 cell lines from advanced melanomas or metastases. Moreover, microarray analysis of 55 human lesions showed higher TSP‐1 expression in primary melanomas and metastases than in common and dysplastic nevi. In a functional enrichment analysis, the expression of TSP‐1 correlated with motility‐related genes. Accordingly, TSP‐1 production was associated with melanoma cell motility in vitro and lung colonization potential in vivo. VEGF/VEGFR‐1 and FGF‐2, involved in melanoma progression, regulated TSP‐1 production. These factors were coexpressed with TSP‐1 and correlated negatively with Slug (SNAI2), a cell migration master gene implicated in melanoma metastasis. We conclude that TSP‐1 cooperates with FGF‐2 and VEGF/VEGFR‐1 in determining melanoma invasion and metastasis, as part of a Slug‐independent motility program.     

Global Invader Impact Network (GIIN): toward standardized evaluation of the ecological impacts of invasive plants

Terrestrial invasive plants are a global problem and are becoming ubiquitous components of most ecosystems. They are implicated in altering disturbance regimes, reducing biodiversity, and changing ecosystem function, sometimes in profound and irreversible ways. However, the ecological impacts of most invasive plants have not been studied experimentally, and most research to date focuses on few types of impacts, which can vary greatly among studies. Thus, our knowledge of existing ecological impacts ascribed to invasive plants is surprisingly limited in both breadth and depth. Our aim was to propose a standard methodology for quantifying baseline ecological impact that, in theory, is scalable to any terrestrial plant invader (e.g., annual grasses to trees) and any invaded system (e.g., grassland to forest). The Global Invader Impact Network (GIIN) is a coordinated distributed experiment composed of an observational and manipulative methodology. The protocol consists of a series of plots located in (1) an invaded area; (2) an adjacent removal treatment within the invaded area; and (3) a spatially separate uninvaded area thought to be similar to pre-invasion conditions of the invaded area. A standardized and inexpensive suite of community, soil, and ecosystem metrics are collected allowing broad comparisons among measurements, populations, and species. The method allows for one-time comparisons and for long-term monitoring enabling one to derive information about change due to invasion over time. Invader removal plots will also allow for quantification of legacy effects and their return rates, which will be monitored for several years. GIIN uses a nested hierarchical scale approach encompassing multiple sites, regions, and continents. Currently, GIIN has network members in six countries, with new members encouraged. To date, study species include representatives of annual and perennial grasses; annual and perennial forbs; shrubs; and trees. The goal of the GIIN framework is to create a standard yet flexible platform for understanding the ecological impacts of invasive plants, allowing both individual and synthetic analyses across a range of taxa and ecosystems. If broadly adopted, this standard approach will offer unique insight into the ecological impacts of invasive plants at local, regional, and global scales.     

Boundary region between coexisting lipid phases as initial binding sites for Escherichia coli alpha-hemolysin: A real-time study

α-Hemolysin (HlyA) is a protein toxin, a member of the pore-forming Repeat in Toxin (RTX) family, secreted by some pathogenic strands of Escherichia coli. The mechanism of action of this toxin seems to involve three stages that ultimately lead to cell lysis: binding, insertion, and oligomerization of the toxin within the membrane. Since the influence of phase segregation on HlyA binding and insertion in lipid membranes is not clearly understood, we explored at the meso- and nanoscale—both in situ and in real-time—the interaction of HlyA with lipid monolayers and bilayers. Our results demonstrate that HlyA could insert into monolayers of dioleoylphosphatidylcholine/sphingomyelin/cholesterol (DOPC/16:0SM/Cho) and DOPC/24:1SM/Cho. The time course for HlyA insertion was similar in both lipidic mixtures. HlyA insertion into DOPC/16:0SM/Cho monolayers, visualized by Brewster-angle microscopy (BAM), suggest an integration of the toxin into both the liquid-ordered and liquid-expanded phases. Atomic-force-microscopy imaging reported that phase boundaries favor the initial binding of the toxin, whereas after a longer time period the HlyA becomes localized into the liquid-disordered (Ld) phases of supported planar bilayers composed of DOPC/16:0SM/Cho. Our AFM images, however, showed that the HlyA interaction does not appear to match the general strategy described for other invasive proteins. We discuss these results in terms of the mechanism of action of HlyA.     

The agro-ecological suitability of Atriplex nummularia and A. halimus for biomass production in Argentine saline drylands

The choice of the best species to cultivate in semi-arid and arid climates is of fundamental importance, and is determined by many factors, including temperature and rainfall, soil type, water availability for irrigation and crop purposes. Soil or water salinity represents one of the major causes of crop stress. Species of the genus Atriplex are characterized by high biomass productivity, high tolerance to drought and salinity, and high efficiency in use of solar radiation and water. Based on a search of the international literature, the authors outline an agro-climatic zoning model to determine potential production areas in Argentina for Atriplex halimus and Atriplex numularia. Using the agroclimatic limits presented in this work, this model may be applied to any part of the world. When superimposed on the saline areas map, the agroclimatic map shows the suitability of agro-ecological zoning for both species for energy purposes on land unsuitable for food production. This innovative study was based on the implementation of a geographic information system that can be updated by further incorporation of complementary information, with consequent improvement of the original database.     

A Highlights from MBoC Selection: Golgi Partitioning Controls Mitotic Entry through Aurora-A Kinase

At the onset of mitosis, the Golgi complex undergoes a multistep fragmentation process that is required for its correct partitioning into the daughter cells. Inhibition of this Golgi fragmentation results in cell cycle arrest at the G2 stage, suggesting that correct inheritance of the Golgi complex is monitored by a “Golgi mitotic checkpoint.” However, the molecular basis of this G2 block is not known. Here, we show that the G2-specific Golgi fragmentation stage is concomitant with centrosome recruitment and activation of the mitotic kinase Aurora-A, an essential regulator for entry into mitosis. We show that a block of Golgi partitioning impairs centrosome recruitment and activation of Aurora-A, which results in the G2 block of cell cycle progression. Overexpression of Aurora-A overrides this cell cycle block, indicating that Aurora-A is a major effector of the Golgi checkpoint. Our findings provide the basis for further understanding of the signaling pathways that coordinate organelle inheritance and cell duplication.     

Direct molecular profiling of minicircle signatures and lineages of Trypanosoma cruzi bloodstream populations causing congenital Chagas disease

Congenital transmission of Trypanosoma cruzi may occur in some or all the gestations from a T. cruzi-infected mother. Variable rates of congenital transmission have been reported in different geographical areas where different parasitic strains predominate, suggesting that parasitic genotypes might play a role in the risk of congenital transmission. Moreover, in cases of transmission it is unknown if the whole maternal T. cruzi population or certain clones are preferentially transmitted by the transplacental route. In this study, bloodstream T. cruzi lineages were identified in blood samples from congenitally infected children, transmitting and non-transmitting mothers and unrelated Chagas disease patients, using improved PCR strategies targeted to nuclear genomic markers. T. cruzi IId was the prevalent genotype among 36/38 PCR-positive congenitally infected infants, 5/5 mothers who transmitted congenital Chagas disease, 12/13 mothers who delivered non-infected children and 28/34 unrelated Chagas disease patients, all coming from endemic localities of Argentina and Bolivia. These figures indicate no association between a particular genotype and vertical transmission. Furthermore, minicircle signatures from the maternal and infants' bloodstream trypanosomes were profiled by restriction fragment length polymorphism of the 330-bp PCR-amplified variable regions in seven cases of mothers and congenitally infected infants. Minicircle signatures were nearly identical between each mother and her infant/s and unique to each mother-infant/s case, a feature that was also observed in twin deliveries. Moreover, allelic size polymorphism analysis of microsatellite loci from populations transmitted to twins showed that all clones from the maternal polyclonal population were equally infective to both siblings.