72 research outputs found
Protocol to train a support vector machine for the automatic curation of bacterial cell detections in microscopy images
No full textSummary: Manual curation of bacterial cell detections in microscopy images remains a time-consuming and laborious task. This work offers a comprehensive, step-by-step tutorial on training a support vector machine to autonomously distinguish between good and bad cell detections. Jupyter notebooks are included to perform feature extraction, labeling, and training of the machine learning model. This method can readily be incorporated into profiling pipelines aimed at extracting a multitude of features across large collections of individual cells, strains, and species.For complete details on the use and execution of this protocol, please refer to Govers et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics
Measuring centimeter-scale sand ripples using multibeam echosounder backscatter data from the Brown Bank area of the Dutch continental shelf
No full textBackscatter data from multibeam echosounders are commonly used to classify seafloor sediment composition. Previously, it was found that the survey azimuth affects backscatter when small organized seafloor structures, such as sand ripples, are present. These sand ripples are too small to be detected in the multibeam bathymetry. Here, we show that such azimuth effects are time dependent and are useful to examine the orientation of sand ripples in relation to the flow direction of the tide. To this end, multibeam echosounder data at four different frequencies were gathered from the area of the Brown Bank in the North Sea. The acoustic results were compared to video and tide-flow data for validation. The sand ripples affected the backscatter at all frequencies, but for the lowest frequencies the effect was spread over more beam angles. Using the acoustic data made it possible to deduce the orientations of the sand ripples over areas of multiple square kilometers. We found that the top centimeter(s) of the seafloor undergoes a complete transformation every six hours, as the orientation of the sand ripples changes with the changing tide. Our methodology allows for morphology change detection at larger scales and higher resolutions than previously achieved
Bimodal Expression of the <i>Salmonella</i> Typhimurium <i>spv</i> Operon
No full textThe well-studied spv operon of Salmonellatyphimurium is important for causing full virulence in mice and both the regulation and function of the Spv proteins have been characterized extensively over the past several decades. Using quantitative single-cell fluorescence microscopy, we demonstrate the spv regulon to display a bimodal expression pattern that originates in the bimodal expression of the SpvR activator. The spv expression pattern is influenced by growth conditions and the specific Styphimurium strain used, but does not require Salmonella-specific virulence regulators. By monitoring real-time promoter kinetics, we reveal that SpvA has the ability to impart negative feedback on spvABCD expression without affecting spvR expression. Together, our data suggest that the SpvA protein counteracts the positive feedback loop imposed by SpvR, and could thus be responsible for dampening spvABCD expression and coordinating virulence protein production in time. The results presented here yield new insights in the intriguing regulation of the spv operon and adds this operon to the growing list of virulence factors exhibiting marked expression heterogeneity in Styphimurium.sponsorship: The authors would like to thank William Cenens, Peter Goos, Kristof Vanoirbeek, Catherine Royer, and Marjan van der Woude for helpful suggestions and fruitful discussions; Sander Van Dromme for helping to screen the Tn5-mVenus mutant library; Victor de Lorenzo for his kind gift of the pBAM1-GFP plasmid; and Johan Paulsson for his kind gift of the pDHL1029 plasmid. This work was supported by doctoral fellowships from the Flemish Agency for Innovation by Science and Technology (IWT-Vlaanderen; to I.P. and S.K.G.) and the Research Foundation - Flanders (FWO-Vlaanderen; to A.C.), and a grant from the Katholieke Universiteit Leuven Research Fund (IDO/10/012). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors declare no conflicts of interest. (Flemish Agency for Innovation by Science and Technology (IWT-Vlaanderen), Research Foundation - Flanders (FWO-Vlaanderen), Katholieke Universiteit Leuven Research Fund|IDO/10/012)status: Publishe
Impact of high hydrostatic pressure on bacterial proteostasis
No full textHigh hydrostatic pressure (HHP) is an important factor that limits microbial growth in deep-sea ecosystems to specifically adapted piezophiles. Furthermore, HHP treatment is used as a novel food preservation technique because of its ability to inactivate pathogenic and spoilage bacteria while minimizing the loss of food quality. Disruption of protein homeostasis (i.e. proteostasis) as a result of HHP-induced conformational changes in ribosomes and proteins has been considered as one of the limiting factors for both microbial growth and survival under HHP conditions. This work therefore reviews the effects of sublethal (≤100MPa) and lethal (>100MPa) pressures on protein synthesis, structure, and functionality in bacteria. Furthermore, current understanding on the mechanisms adopted by piezophiles to maintain proteostasis in HHP environments and responses developed by atmospheric-adapted bacteria to protect or restore proteostasis after HHP exposure are discussed.sponsorship: This work was supported by a doctoral fellowship of the Flemish Agency for Innovation by Science and Technology (IWT-Vlaanderen to S.K.G.), by a postdoctoral fellowship of the Research Foundation of Flanders (FWO-Vlaanderen to E.G.), and by grants of the Research Foundation of Flanders (FWO-Vlaanderen; grant G.0580.11) and the KU Leuven Research Fund (IDO/10/012 and STRT1/10/036). (Flemish Agency for Innovation by Science and Technology (IWT-Vlaanderen), Research Foundation of Flanders (FWO-Vlaanderen), Research Foundation of Flanders (FWO-Vlaanderen)|G.0580.11, KU Leuven Research Fund|IDO/10/012, KU Leuven Research Fund|STRT1/10/036)status: Publishe
Design capability in a software SME: Report from an embedded design innovation catalyst
No full textRecent studies on the added value that design provides to firms has led to widespread interest amongst the business community to develop design. However, knowledge of how organizations of varying industry types actually make use of design to generate competitive advantages remains limited. This study investigates a small- to medium-sized enterprise (SME) software firm that had recently taken steps to develop design capability1. The Dutch firm, titled CM, identified a need to increase design capability as a source of added competitiveness. During a six-month period, a design innovation catalyst (DIC) was embedded in the firm to build and integrate design capability across the firm. During the study’s duration, the catalyst found the barriers to design capability to be a prevailing data-driven approach to value creation, reliance on self-referential knowledge rather than hypothesis testing with customers and users and a general low urgency to embrace design. The manner in which CM now leverages design as a value creation mechanism is shaped by addressing the opposing barriers to change that were encountered within the firm. This paper contributes practical knowledge on how design can be built quickly over six months and become a vehicle for a software firm to move from data-driven to user-centred solutions. Marketing and Consumer Researc
Seafloor characterization using multibeam echosounder backscatter data: Methodology and results in the north sea
No full textSeafloor characterization using multibeam echosounder (MBES) backscatter data is an active field of research. The observed backscatter curve (OBC) is used in an inversion algorithm with available physics-based models to determine the seafloor geoacoustic parameters. A complication is that the OBC cannot directly be coupled to the modeled backscatter curve (MBC) due to the correction of uncalibrated sonars. Grab samples at reference areas are usually required to estimate the angular calibration curve (ACC) prior to the inversion. We first attempt to estimate the MBES ACC without grab sampling by using the least squares cubic spline approximation method implemented in a differential evolution optimization algorithm. The geoacoustic parameters are then inverted over the entire area using the OBCs corrected for the estimated ACC. The results indicate that a search for at least three geoacoustic parameters is required, which includes the sediment mean grain size, roughness parameter, and volume scattering parameter. The inverted mean grain sizes are in agreement with grab samples, indicating reliability and stability of the proposed method. Furthermore, the interaction between the geoacoustic parameters and Bayesian acoustic classes is investigated. It is observed that higher backscatter values, and thereby higher acoustic classes, should not only be attributed to (slightly) coarser sediment, especially in a homogeneous sedimentary environment such as the Brown Bank, North Sea. Higher acoustic classes should also be attributed to larger seafloor roughness and volume scattering parameters, which are not likely intrinsic to only sediment characteristics but also to other contributing factors.Aircraft Noise and Climate Effect
Label-induced mislocalization of IbpA in exponential phase <i>E</i>. <i>coli</i> cells.
No full text(A) Representative phase contrast, epifluorescence (reporting IbpA expression/production and localization), and superimposed images of E. coli MG1655 cells containing the indicated IbpA fluorescent fusion proteins. Scale bars correspond to 2 μm. (B) Measured distribution of punctate and diffuse fluorescence intensity for the indicated fusion proteins. The fluorescence intensity distribution of at least 25 individual cells was determined per independent experiment. (C) The average number of observed foci per cell for the indicated fusion proteins. Per independent experiment, at least 100 cells were examined to determine the average number of cellular foci. ND = no foci could be detected. For both (B) and (C), the means of 3 independent experiments are shown, with error bars representing the standard deviation between experiments. (D) Cumulative distribution of average cellular mCherry fluorescence of MG1655 pSG1, MG1655 ibpA-yfp pSG1, and MG1655 ibpA-msfgfp pSG1 cells, derived from 3 independent experiments (n ≥ 80 cells per independent experiment). pSG1 is a low-copy plasmid containing a transcriptional PibpA-mCherry reporter. A K-S test (p-value = 3.97 × 10−19) indicated significantly increased PibpA expression in MG1655 ibpA-yfp cells, whereas this was not the case for MG1655 ibpA-msfgfp cells (K-S test; p-value = 0.15). (E) Histograms showing the distribution of average cellular GFP fluorescence of control and heat-treated (47 °C, 15 min) MG1655 ibpA-msfgfp cells, derived from 3 independent experiments (n ≥ 129 per independent experiment). Elevated IbpA levels could be detected in the heat-treated samples (K-S test; p-value = 2.23 × 10−223). (F) Representative phase contrast, GFP epifluorescence (reporting IbpA expression/production and localization), and superimposed images of MG1655 ibpA-msfgfp cells exposed to a sublethal heat shock (47 °C, 15 min). Scale bar corresponds to 2 μm. (G) Binned histograms showing the PA distribution along the relative axial position of MG1655 ibpA-msfgfp cells (n = 641 cells) exposed to a sublethal heat shock (47 °C, 15 min). (H) Heat map displaying the distribution of PA localization in these cells (n = 457 cells). The numerical data underlying this figure can be found in S1 Data. A.U., arbitrary unit; GFP, green fluorescent protein; IbpA, inclusion body binding protein A; K-S, Kolmogorov-Smirnov; mCer, monomeric cerulean; msfGFP, monomeric superfolder GFP; PA, protein aggregate; PibpA, ibpA promoter; YFP, yellow fluorescent protein.</p
Severely heat injured survivors of E. coli O157:H7 ATCC 43888 display variable and heterogeneous stress resistance behavior
No full textAlthough minimal food processing strategies aim to eliminate foodborne pathogens and spoilage microorganisms through a combination of mild preservation techniques, little is actually known on the resistance behavior of the small fraction of microorganisms surviving an inimical treatment. In this study, the conduct of severely heat stressed survivors of E. coli O157:H7 ATCC 43888, as an indicator for the low infectious dose foodborne enterohaemorrhagic strains, was examined throughout their resuscitation and outgrowth. Despite the fact that these survivors were initially sublethally injured, they were only marginally more sensitive to a subsequent heat treatment and actually much more resistant to a subsequent high hydrostatic pressure (HHP) shock in comparison with unstressed control cells. Throughout further resuscitation, however, their initial HHP resistance rapidly faded out, while their heat resistance increased and surpassed the initial heat resistance of unstressed control cells. Results also indicated that the population eventually emerging from the severely heat stressed survivors heterogeneously consisted of both growing and non-growing persister-like cells. Together, these observations provide deeper insights into the particular behavior and heterogeneity of stressed foodborne pathogens in the context of food preservation
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