Functional Identification of Conserved Residues Involved in Lactobacillus rhamnosus Strain GG Sortase Specificity and Pilus Biogenesis

In Gram-positive bacteria, sortase-dependent pili mediate the adhesion of bacteria to host epithelial cells and play a pivotal role in colonization, host signaling, and biofilm formation. Lactobacillus rhamnosus strain GG, a well known probiotic bacterium, also displays on its cell surface mucus-binding pilus structures, along with other LPXTG surface proteins, which are processed by sortases upon specific recognition of a highly conserved LPXTG motif. Bioinformatic analysis of all predicted LPXTG proteins encoded by the L. rhamnosus GG genome revealed a remarkable conservation of glycine residues juxtaposed to the canonical LPXTG motif. Here, we investigated and defined the role of this so-called triple glycine (TG) motif in determining sortase specificity during the pilus assembly and anchoring. Mutagenesis of the TG motif resulted in a lack or an alteration of the L. rhamnosus GG pilus structures, indicating that the TG motif is critical in pilus assembly and that they govern the pilin-specific and housekeeping sortase specificity. This allowed us to propose a regulatory model of the L. rhamnosus GG pilus biogenesis. Remarkably, the TG motif was identified in multiple pilus gene clusters of other Gram-positive bacteria, suggesting that similar signaling mechanisms occur in other, mainly pathogenic, species.     

A Push-Pull CORF Model of a Simple Cell with Antiphase Inhibition Improves SNR and Contour Detection

We propose a computational model of a simple cell with push-pull inhibition, a property that is observed in many real simple cells. It is based on an existing model called Combination of Receptive Fields or CORF for brevity. A CORF model uses as afferent inputs the responses of model LGN cells with appropriately aligned center-surround receptive fields, and combines their output with a weighted geometric mean. The output of the proposed model simple cell with push-pull inhibition, which we call push-pull CORF, is computed as the response of a CORF model cell that is selective for a stimulus with preferred orientation and preferred contrast minus a fraction of the response of a CORF model cell that responds to the same stimulus but of opposite contrast. We demonstrate that the proposed push-pull CORF model improves signal-to-noise ratio (SNR) and achieves further properties that are observed in real simple cells, namely separability of spatial frequency and orientation as well as contrast-dependent changes in spatial frequency tuning. We also demonstrate the effectiveness of the proposed push-pull CORF model in contour detection, which is believed to be the primary biological role of simple cells. We use the RuG (40 images) and Berkeley (500 images) benchmark data sets of images with natural scenes and show that the proposed model outperforms, with very high statistical significance, the basic CORF model without inhibition, Gabor-based models with isotropic surround inhibition, and the Canny edge detector. The push-pull CORF model that we propose is a contribution to a better understanding of how visual information is processed in the brain as it provides the ability to reproduce a wider range of properties exhibited by real simple cells. As a result of push-pull inhibition a CORF model exhibits an improved SNR, which is the reason for a more effective contour detection.     

Inhibition of Aurora Kinase B Is Important for Biologic Activity of the Dual Inhibitors of BCR-ABL and Aurora Kinases R763/AS703569 and PHA-739358 in BCR-ABL Transformed Cells

ABL tyrosine kinase inhibitors (TKI) like Imatinib, Dasatinib and Nilotinib are the gold standard in conventional treatment of CML. However, the emergence of resistance remains a major problem. Alternative therapeutic strategies of ABL TKI-resistant CML are urgently needed. We asked whether dual inhibition of BCR-ABL and Aurora kinases A-C could overcome resistance mediated by ABL kinase mutations. We therefore tested the dual ABL and Aurora kinase inhibitors PHA-739358 and R763/AS703569 in Ba/F3- cells ectopically expressing wild type (wt) or TKI-resistant BCR-ABL mutants. We show that both compounds exhibited strong anti-proliferative and pro-apoptotic activity in ABL TKI resistant cell lines including cells expressing the strongly resistant T315I mutation. Cell cycle analysis indicated polyploidisation, a consequence of continued cell cycle progression in the absence of cell division by Aurora kinase inhibition. Experiments using drug resistant variants of Aurora B indicated that PHA-739358 acts on both, BCR-ABL and Aurora Kinase B, whereas Aurora kinase B inhibition might be sufficient for the anti-proliferative activity observed with R763/AS703569. Taken together, our data demonstrate that dual ABL and Aurora kinase inhibition might be used to overcome ABL TKI resistant CML.     

Climate-based models for understanding and forecasting dengue epidemics

Background

Dengue dynamics are driven by complex interactions between human-hosts, mosquito-vectors and viruses that are influenced by environmental and climatic factors. The objectives of this study were to analyze and model the relationships between climate, Aedes aegypti vectors and dengue outbreaks in Noumea (New Caledonia), and to provide an early warning system.

Methodology/Principal Findings

Epidemiological and meteorological data were analyzed from 1971 to 2010 in Noumea. Entomological surveillance indices were available from March 2000 to December 2009. During epidemic years, the distribution of dengue cases was highly seasonal. The epidemic peak (March–April) lagged the warmest temperature by 1–2 months and was in phase with maximum precipitations, relative humidity and entomological indices. Significant inter-annual correlations were observed between the risk of outbreak and summertime temperature, precipitations or relative humidity but not ENSO. Climate-based multivariate non-linear models were developed to estimate the yearly risk of dengue outbreak in Noumea. The best explicative meteorological variables were the number of days with maximal temperature exceeding 32°C during January–February–March and the number of days with maximal relative humidity exceeding 95% during January. The best predictive variables were the maximal temperature in December and maximal relative humidity during October–November–December of the previous year. For a probability of dengue outbreak above 65% in leave-one-out cross validation, the explicative model predicted 94% of the epidemic years and 79% of the non epidemic years, and the predictive model 79% and 65%, respectively.

Conclusions/Significance

The epidemic dynamics of dengue in Noumea were essentially driven by climate during the last forty years. Specific conditions based on maximal temperature and relative humidity thresholds were determinant in outbreaks occurrence. Their persistence was also crucial. An operational model that will enable health authorities to anticipate the outbreak risk was successfully developed. Similar models may be developed to improve dengue management in other countries.     

Direct behavioral evidence for retronasal olfaction in rats

The neuroscience of flavor perception is becoming increasingly important to understand abnormal feeding behaviors and associated chronic diseases such as obesity. Yet, flavor research has mainly depended on human subjects due to the lack of an animal model. A crucial step towards establishing an animal model of flavor research is to determine whether the animal uses the retronasal mode of olfaction, an essential element of flavor perception. We designed a go- no go behavioral task to test the rat's ability to detect and discriminate retronasal odorants. In this paradigm, tasteless aqueous solutions of odorants were licked by water-restricted head-fixed rats from a lick spout. Orthonasal contamination was avoided by employing a combination of a vacuum around the lick-spout and blowing clean air toward the nose. Flow models support the effectiveness of both approaches. The licked odorants were successfully discriminated by rats. Moreover, the tasteless odorant amyl acetate was reliably discriminated against pure distilled water in a concentration-dependent manner. The results from this retronasal odor discrimination task suggest that rats are capable of smelling retronasally. This direct behavioral evidence establishes the rat as a useful animal model for flavor research.     

Characterization of the SpaCBA pilus fibers in the probiotic Lactobacillus rhamnosus GG

Lactobacillus rhamnosus GG is a human intestinal isolate that has been studied intensively because of its probiotic properties. We have previously shown that L. rhamnosus GG produces proteinaceous pili that earlier had been observed only in Gram-positive pathogens (M. Kankainen et al., Proc. Natl. Acad. Sci. U. S. A. 106:17193-17198, 2009). These pili were found to be encoded by the spaCBA gene cluster, and the pilus-associated SpaC pilin was shown to confer on the cells a mucus-binding ability. In addition to the spaCBA cluster, another putative pilus cluster, spaFED, was predicted from the L. rhamnosus GG genome sequence. Herein, we show that only SpaCBA pili are produced by L. rhamnosus, and we describe a detailed analysis of cell wall-associated and affinity-purified SpaCBA pili by Western blotting and immunogold electron microscopy. Our results indicate that SpaCBA pili are heterotrimeric protrusions with a SpaA subunit as the shaft-forming major pilin. Only a few SpaB subunits could be observed in pilus fibers. Instead, SpaB pilins were found at pilus bases, as assessed by immunogold double labeling of thin sections of cells, suggesting that SpaB is involved in the termination of pilus assembly. The SpaC adhesin was present along the whole pilus length at numbers nearly equaling those of SpaA. The relative amount and uniform distribution of SpaC within pili not only makes it possible to exert both long-distance and intimate contact with host tissue but also provides mucus-binding strength, which explains the prolonged intestinal residency times observed for L. rhamnosus GG compared to that of nonpiliated lactobacilli.     

Effects of aging and arm swing on the metabolic cost of stability in human walking

To gain insight into the mechanical determinants of walking energetics, we investigated the effects of aging and arm swing on the metabolic cost of stabilization. We tested two hypotheses: (1) elderly adults consume more metabolic energy during walking than young adults because they consume more metabolic energy for lateral stabilization, and (2) arm swing reduces the metabolic cost of stabilization during walking in young and elderly adults. To test these hypotheses, we provided external lateral stabilization by applying bilateral forces (10% body weight) to a waist belt via elastic cords while young and elderly subjects walked at 1.3m/s on a motorized treadmill with arm swing and with no arm swing. We found that the external stabilizer reduced the net rate of metabolic energy consumption to a similar extent in elderly and young subjects. This reduction was greater (6-7%) when subjects walked with no arm swing than when they walked normally (3-4%). When young or elderly subjects eliminated arm swing while walking with no external stabilization, net metabolic power increased by 5-6%. We conclude that the greater metabolic cost of walking in elderly adults is not caused by a greater cost of lateral stabilization. Moreover, arm swing reduces the metabolic cost of walking in both young and elderly adults likely by contributing to stability.     

Serotyping of Toxoplasma gondii: striking homogeneous pattern between symptomatic and asymptomatic infections within Europe and South America

Field isolates of Toxoplasma gondii in Europe and North America have been grouped into three clonal lineages that display different virulence in mice. Whether the genetic structure of the parasite is related to clinical expression in humans has not yet been demonstrated. We developed an enzyme-linked immunosorbent assay which uses lineage-specific, polymorphic polypeptides derived from the dense granule antigens, GRA5 and GRA6. Our goal was to compare serotypical patterns observed in asymptomatic versus symptomatic (ocular disease and severe infection in human immunodeficiency virus (HIV)-positive patients) infections among patients from Europe and South America. Independent of the clinical presentation of the disease, serotypes differed according to geographical origin, with a homogeneous distribution of serotype II in Europe and of serotypes I and III in South America. We conclude that GRA5-GRA6 serotyping is an interesting tool to study serotype prevalence in populations but it is not an accurate marker of pathogenicity of Toxoplasma infection in humans.     

Interactive effects of global and regional change on a coastal ecosystem

Vertical distribution and diel vertical migration of a zooplankton community were studied at two stations off Central Peru in April 2006. Zooplankton was collected at five depth strata by vertical hauls with Hydo-Bios multinet (300-μm mesh, 0.25-m² mouth size). The zooplankton community was distributed in relation to a strong, shallow oxycline (1 ml l^?1 oxygen isopleth generally above 36 m). The highest total abundance was always in the upper, well-oxygenated layer. The most important species were: Acartia tonsa (72.86%), Centropages brachiatus (7.5%), and Paracalanus parvus (3.1%); Acartia tonsa was the dominant species at all times. Larvae of the polychaete Magelona sp. (7.5%) and larvae of the brachiopod Discinisca lamellosa (3.5%) were numerically dominant in April and small copepods e.g. Oncaea venusta (3.88%) were numerically dominant during August. Five distinct patterns of vertical distribution and migration in relation to the oxygen minimum layer were distinguished in this study: (1) Ontogenetic vertical migration through the oxycline (Acartia tonsa adults, nauplii, and copepodids), (2) permanent limitation to layers above the oxycline (e.g. Oikopleura sp., most invertebrate larvae), (3) distribution mostly below the oxycline with occasional migration into the layers just above the oxycline (Eucalanus inermis), (4) Diel Vertical Migration (Centropages brachiatus), and (5) reverse Diel Vertical Migration (larvae of the polychaete Magelona sp.).     

Land use types influenced avian assemblage structure in a forest–agriculture landscape in Ghana

The conservation of biodiversity within tropical forest regions does not lie only in the maintenance of natural forest areas, but on conservation strategies directed toward agricultural land types within which they are embedded. This study investigated variations in bird assemblages of different functional groups of forest‐dependent birds in three agricultural land types, relative to distance from the interior of 34 tropical forest patches of varying sizes. Point counts were used to sample birds at each study site visited. Data from counts were used to estimate species richness, species evenness, and Simpson's diversity of birds. Mean species richness, evenness, and diversity were modeled as responses and as a function of agricultural land type, distance from the forest interior and three site‐scale vegetation covariates (density of large trees, fruiting trees, and patch size) using generalized linear mixed‐effect models. Mean observed species richness of birds varied significantly within habitat types. Mean observed species richness was highest in forest interior sites while sites located in farm centers recorded the lowest mean species richness. Species richness of forest specialists was strongly influenced by the type of agricultural land use. Fallow lands, density of large trees, and patch size strongly positively influenced forest specialists. Insectivorous and frugivorous birds were more species‐rich in fallow lands while monoculture plantations favored nectarivorous birds. Our results suggest that poor agricultural practices can lead to population declines of forest‐dependent birds particularly specialist species. Conservation actions should include proper land use management that ensures heterogeneity through retention of native tree species on farms in tropical forest‐agriculture landscapes.