Explore the vast range of titles available.

Forums and email lists play a major role in assisting scientists in using software. Previously, each open-source bioimaging software package had its own distinct forum or email list. Although each provided access to experts from various software teams, this fragmentation resulted in many scientists not knowing where to begin with their projects. Thus, the scientific imaging community lacked a central platform where solutions could be discussed in an open, software-independent manner. In response, we introduce the Scientific Community Image Forum, where users can pose software-related questions about digital image analysis, acquisition, and data management.

Background ImageJ is an image analysis program extensively used in the biological sciences and beyond. Due to its ease of use, recordable macro language, and extensible plug-in architecture, ImageJ enjoys contributions from non-programmers, amateur programmers, and professional developers alike. Enabling such a diversity of contributors has resulted in a large community that spans the biological and physical sciences. However, a rapidly growing user base, diverging plugin suites, and technical limitations have revealed a clear need for a concerted software engineering effort to support emerging imaging paradigms, to ensure the software’s ability to handle the requirements of modern science. Results We rewrote the entire ImageJ codebase, engineering a redesigned plugin mechanism intended to facilitate extensibility at every level, with the goal of creating a more powerful tool that continues to serve the existing community while addressing a wider range of scientific requirements. This next-generation ImageJ, called “ImageJ2” in places where the distinction matters, provides a host of new functionality. It separates concerns, fully decoupling the data model from the user interface. It emphasizes integration with external applications to maximize interoperability. Its robust new plugin framework allows everything from image formats, to scripting languages, to visualization to be extended by the community. The redesigned data model supports arbitrarily large, N-dimensional datasets, which are increasingly common in modern image acquisition. Despite the scope of these changes, backwards compatibility is maintained such that this new functionality can be seamlessly integrated with the classic ImageJ interface, allowing users and developers to migrate to these new methods at their own pace. Conclusions Scientific imaging benefits from open-source programs that advance new method development and deployment to a diverse audience. ImageJ has continuously evolved with this idea in mind; however, new and emerging scientific requirements have posed corresponding challenges for ImageJ’s development. The described improvements provide a framework engineered for flexibility, intended to support these requirements as well as accommodate future needs. Future efforts will focus on implementing new algorithms in this framework and expanding collaborations with other popular scientific software suites.

Significance Shigella spp . are responsible for devastating diarrheal diseases, primarily in children, within underdeveloped countries. Shigella invades the mucosa of the large intestine, causing inflammation and damage to the epithelium. Here, we have measured the progression of Shigella infection in a small animal model of the disease to better understand the mechanism of invasion of the colonic mucosa. The novelty of our approach relies on the tracking of fluorescent bacteria inside the infected tissue at various time points using confocal microscopy and subsequent quantitative bioimage analyses. Our approach is readily applicable to other host–pathogen systems to quantify host–pathogen interactions. Few studies within the pathogenic field have used advanced imaging and analytical tools to quantitatively measure pathogenicity in vivo. In this work, we present a novel approach for the investigation of host–pathogen processes based on medium-throughput 3D fluorescence imaging. The guinea pig model for Shigella flexneri invasion of the colonic mucosa was used to monitor the infectious process over time with GFP-expressing S. flexneri . A precise quantitative imaging protocol was devised to follow individual S. flexneri in a large tissue volume. An extensive dataset of confocal images was obtained and processed to extract specific quantitative information regarding the progression of S. flexneri infection in an unbiased and exhaustive manner. Specific parameters included the analysis of S. flexneri positions relative to the epithelial surface, S. flexneri density within the tissue, and volume of tissue destruction. In particular, at early time points, there was a clear association of S. flexneri with crypts, key morphological features of the colonic mucosa. Numerical simulations based on random bacterial entry confirmed the bias of experimentally measured S. flexneri for early crypt targeting. The application of a correlative light and electron microscopy technique adapted for thick tissue samples further confirmed the location of S. flexneri within colonocytes at the mouth of crypts. This quantitative imaging approach is a novel means to examine host–pathogen systems in a tailored and robust manner, inclusive of the infectious agent.

The specific contribution of each bacterial species within a complex microbiota to the regulation of host lipid metabolism remains largely unknown. Using two model commensal microorganisms, L. paracasei and E. coli , we demonstrated that both bacterial species impacted host lipid metabolism in a diet-dependent manner and, notably, that L. paracasei -colonized mice but not E. coli -colonized mice resisted high-fat-diet-induced body weight gain. In addition, we set up cellular models of fatty acid absorption and secretion by enterocytes cocultured with bacteria and showed that, in vitro , both L. paracasei and E. coli inhibited lipid secretion, through increased intracellular fat storage and enhanced lipid catabolism, respectively. The gut microbiota contributes to nutrients absorption and metabolism by enterocytes, but the molecular mechanisms involved remain poorly understood, and most conclusions are inferred from studies comparing germfree and conventional animals colonized with diverse bacterial species. We selected two model commensal microorganisms, Escherichia coli and Lactobacillus paracasei , to assess the role of the small-intestinal microbiota in modulating lipid absorption and metabolism by the epithelium. Using an integrated approach encompassing cellular and murine models and combining metabolic parameters measurement, lipid droplet imaging, and gene expression analysis, we demonstrated that under homeostatic conditions, L. paracasei promotes fat storage in enterocytes, whereas E. coli enhances lipid catabolism and reduces chylomicron circulating levels. The Akt/mammalian target of sirolimus (mTOR) pathway is inhibited by both bacterial species in vitro , indicating that several regulatory pathways are involved in the distinct intracellular lipid outcomes associated with each bacterial species. Moreover, soluble bacterial factors partially reproduce the effects observed with live microorganisms. However, reduction of chylomicron circulating levels in E. coli -colonized animals is lost under high-fat-diet conditions, whereas it is potentiated by L. paracasei colonization accompanied by resistance to hypercholesterolemia and excess body weight gain. IMPORTANCE The specific contribution of each bacterial species within a complex microbiota to the regulation of host lipid metabolism remains largely unknown. Using two model commensal microorganisms, L. paracasei and E. coli , we demonstrated that both bacterial species impacted host lipid metabolism in a diet-dependent manner and, notably, that L. paracasei -colonized mice but not E. coli -colonized mice resisted high-fat-diet-induced body weight gain. In addition, we set up cellular models of fatty acid absorption and secretion by enterocytes cocultured with bacteria and showed that, in vitro , both L. paracasei and E. coli inhibited lipid secretion, through increased intracellular fat storage and enhanced lipid catabolism, respectively.

The gallbladder is often colonized by Salmonella during typhoid fever, yet little is known about bacterial pathogenesis in this organ. With use of a mouse model of acute typhoid fever, we demonstrate that Salmonella infect gallbladder epithelial cells in vivo. Bacteria in the gallbladder showed a unique behavior as they replicated within gallbladder epithelial cells and remained confined to those cells without translocating to the mucosa. Infected gallbladders showed histopathological damage characterized by destruction of the epithelium and massive infiltration of neutrophils, accompanied by a local increase of proinflammatory cytokines. Damage was determined by the ability of Salmonella to invade gallbladder epithelial cells and was independent of high numbers of replication-competent, although invasion-deficient, bacteria in the lumen. Our results establish gallbladder epithelial cells as a novel niche for in vivo replication of Salmonella and reveal the involvement of these cells in the pathogenesis of Salmonella in the gallbladder during the course of acute typhoid fever

The Gram-negative enteroinvasive bacterium Shigella flexneri is responsible for the endemic form of bacillary dysentery, an acute rectocolitis in humans. S. flexneri uses a type III secretion system to inject effector proteins into host cells, thus diverting cellular functions to its own benefit. Protective immunity to reinfection requires several rounds of infection to be elicited and is short-lasting, suggesting that S. flexneri interferes with the priming of specific immunity. Considering the key role played by T-lymphocyte trafficking in priming of adaptive immunity, we investigated the impact of S. flexneri on T-cell dynamics in vivo. By using two-photon microscopy to visualize bacterium–T-cell cross-talks in the lymph nodes, where the adaptive immunity is initiated, we provide evidence that S. flexneri , via its type III secretion system, impairs the migration pattern of CD4 ⁺ T cells independently of cognate recognition of bacterial antigens. We show that bacterial invasion of CD4 ⁺ T lymphocytes occurs in vivo, and results in cell migration arrest. In the absence of invasion, CD4 ⁺ T-cell migration parameters are also dramatically altered. Signals resulting from S. flexneri interactions with subcapsular sinus macrophages and dendritic cells, and recruitment of polymorphonuclear cells are likely to contribute to this phenomenon. These findings indicate that S. flexneri targets T lymphocytes in vivo and highlight the role of type III effector secretion in modulating host adaptive immune responses.

Current workflows for colocalization analysis in fluorescence microscopic imaging introduce significant bias in terms of the user’s choice of region of interest (ROI). In this work, we introduce an automatic, unbiased structured detection method for correlated region detection between two random processes observed on a common domain. We argue that although intuitive, using the maximum log-likelihood statistic directly suffers from potential bias and substantially reduced power. Therefore, we introduce a simple size-based normalization to overcome this problem. We show that scanning using the proposed statistic leads to optimal correlated region detection over a large collection of structured correlation detection problems.

ABSTRACT Shigella spp. are bacterial pathogens that invade the human colonic mucosa using a type III secretion apparatus (T3SA), a proteinaceous device activated upon contact with host cells. Active T3SAs translocate proteins that carve the intracellular niche of Shigella spp. Nevertheless, the activation state of the T3SA has not been addressed in vivo. Here, we used a green fluorescent protein transcription-based secretion activity reporter (TSAR) to provide a spatio-temporal description of S. flexneri T3SAs activity in the colon of Guinea pigs. First, we observed that early mucus release is triggered in the vicinity of luminal bacteria with inactive T3SA. Subsequent mucosal invasion showed bacteria with active T3SA associated with the brush border, eventually penetrating into epithelial cells. From 2 to 8 h post-challenge, the infection foci expanded, and these intracellular bacteria displayed homogeneously high-secreting activity, while extracellular foci within the lamina propria featured bacteria with low secretion activity. We also found evidence that within lamina propria macrophages, bacteria reside in vacuoles instead of accessing the cytosol. Finally, bacteria were cleared from tissues between 8 and 24 h post-challenge, highlighting the hit-and-run colonization strategy of Shigella. This study demonstrates how genetically encoded reporters can contribute to deciphering pathogenesis in vivo. We mapped Shigella flexneri cells and assessed the activity of their type III secretion apparatus in the large intestine of Guinea pigs.

Pathogenic enterobacteria face various oxygen (O 2 ) levels during intestinal colonization from the O 2 -deprived lumen to oxygenated tissues. Using Shigella flexneri as a model, we have previously demonstrated that epithelium invasion is promoted by O 2 in a type III secretion system-dependent manner. However, subsequent pathogen adaptation to tissue oxygenation modulation remained unknown. Assessing single-cell distribution, together with tissue oxygenation, we demonstrate here that the colonic mucosa O 2 is actively depleted by S. flexneri aerobic respiration—and not host neutrophils—during infection, leading to the formation of hypoxic foci of infection. This process is promoted by type III secretion system inactivation in infected tissues, favouring colonizers over explorers. We identify the molecular mechanisms supporting infectious hypoxia induction, and demonstrate here how enteropathogens optimize their colonization capacity in relation to their ability to manipulate tissue oxygenation during infection. Shigella flexneri aerobic respiration depletes mucosal oxygen in the colon to promote infection.

Background Enterohepatic bacterial infections have the potential to affect multiple physiological processes of the body. Fibroblast growth factor 15/19 (FGF15 in mice, FGF19 in humans) is a hormone that functions as a central regulator of glucose, lipid and bile acid metabolism. FGF15/19 is produced in the intestine and exert its actions on the liver by signaling through the FGFR4-βKlotho receptor complex. Here, we examined the in vivo effects of enterohepatic bacterial infection over the FGF15 endocrine axis. Results Infection triggered significant reductions in the intestinal expression of Fgf15 and its hepatic receptor components ( Fgfr4 and Klb (βKlotho)). Infection also resulted in alterations of the expression pattern of genes involved in hepatobiliary function, marked reduction in gallbladder bile volumes and accumulation of hepatic cholesterol and triglycerides. The decrease in ileal Fgf15 expression was associated with liver bacterial colonization and hepatobiliary pathophysiology rather than with direct intestinal bacterial pathogenesis. Conclusions Bacterial pathogens of the enterohepatic system can disturb the homeostasis of the FGF15/19-FGFR4 endocrine axis. These results open up a possible link between FGF15/19-FGFR4 disruptions and the metabolic and nutritional disorders observed in infectious diseases.