Tracking the stochastic growth of bacterial populations in microfluidic droplets

Bacterial growth in microfluidic droplets is relevant in biotechnology, in microbial ecology, and in understanding stochastic population dynamics in small populations. However, it has proved challenging to automate measurement of absolute bacterial numbers within droplets, forcing the use of proxy measures for population size. Here we present a microfluidic device and imaging protocol that allows high-resolution imaging of thousands of droplets, such that individual bacteria stay in the focal plane and can be counted automatically. Using this approach, we track the stochastic growth of hundreds of replicate Escherichia coli populations within droplets. We find that, for early times, the statistics of the growth trajectories obey the predictions of the Bellman-Harris model, in which there is no inheritance of division time. Our approach should allow further testing of models for stochastic growth dynamics, as well as contributing to broader applications of droplet-based bacterial culture

Gut overgrowth harms the critically ill patient requiring treatment on the intensive care unit

Overgrowth is defined as ≥105 potential pathogens per ml of saliva and/or per g of faeces. There are six \u27normal\u27 potential pathogens carried by healthy individuals and nine \u27abnormal\u27 potential pathogens carried by individuals with underlying disease both chronic and acute. Surveillance cultures of throat and/or rectum are required to identify overgrowth of \u27normal\u27 and/or \u27abnormal\u27 potential pathogens. There is a qualitative and quantitative relationship between surveillance samples and diagnostic samples of tracheal aspirate and blood, i.e., as soon as potential pathogens reach overgrowth concentrations in the surveillance samples, the diagnostic samples become positive for identical potential pathogens. Digestive tract decontamination aims at the eradication of overgrowth in order to prevent severe infections of lower airways and blood. Parenteral cefotaxime controls overgrowth of \u27normal\u27 bacteria, and enteral polyenes control overgrowth of \u27normal\u27 Candida species. Enteral polymyxin and tobramycin (with or without) vancomycin control \u27abnormal\u27 overgrowth

The addition of enteral to parenteral antimicrobials may prolong antibiotic era

Resistance to parenteral antimicrobials generally occurs within two years after introduction into general use. The site where de novo resistance develops has been acknowledged to be the gut. Overgrowth of abnormal flora, defined as 105 potential pathogens per g of faeces is a risk factor for resistance following increased spontaneous mutation leading to polyclonality and antimicrobial resistance. As parenteral antimicrobials generally fail to eradicate the abnormal carrier state in overgrowth concentrations due to sub-lethal concentrations in bile and mucus the enteral antimicrobials polymyxin/tobramycin aiming at converting the abnormal carrier state into normal carriage, are the essential component of selective decontamination of the digestive tract (SDD), because they eradicate carriage and overgrowth including resistant mutants, maintaining the usefulness of parenteral antimicrobials

Oxygen-enhanced MRI and radiotherapy in patients with oropharyngeal squamous cell carcinoma

Background and purpose: This study aimed to assess the role of T1 mapping and oxygen-enhanced MRI in patients undergoing radical dose radiotherapy for HPV positive oropharyngeal cancer, which has not yet been examined in an OE-MRI study. Materials and methods: Variable Flip Angle T1 maps were acquired on a 3T MRI scanner while patients (n = 12) breathed air and/or 100 % oxygen, before and after fraction 10 of the planned 30 fractions of chemoradiotherapy (‘visit 1’ and ‘visit 2’, respectively). The analysis aimed to assess to what extent (1) native R1 relates to patient outcome; (2) OE-MRI response relates to patient outcome; (3) changes in mean R1 before and after radiotherapy related to clinical outcome in patients with oropharyngeal squamous cell carcinoma. Results: Due to the radiotherapy being largely successful, the sample sizes of non-responder groups were small, and therefore it was not possible to properly assess the predictive nature of OE-MRI. The tumour R1 increased in some patients while decreasing in others, in a pattern that was overall consistent with the underlying OE-MRI theory and previously reported tumour OE-MRI responses. In addition, we discuss some practical challenges faced when integrating this technique into a clinical trial, with the aim that sharing this is helpful to researchers planning to use OE-MRI in future clinical studies. Conclusion: Altogether, these results suggest that further clinical OE-MRI studies to assess hypoxia and radiotherapy response are worth pursuing, and that there is important work to be done to improve the robustness of the OE-MRI technique in human applications in order for it to be useful as a widespread clinical technique

Assessment of ridden horse behavior

Assessments of the behavior of ridden horses form the basis of performance evaluation. The purpose of any performance being evaluated will determine the factors considered important, those indicative of 'poor' performance and what makes a successful equine athlete. Currently there is no consistent objective means of assessing ridden horse behavior and inevitably, given the different equestrian disciplines, the likelihood of a universal standard of good and bad performance is remote. Nevertheless, in order to protect the welfare of the ridden horse regardless of its specific role, we should strive for consensus on an objective means of identifying behavioral signs indicative of mental state. Current technological developments enable objective evaluation of movement patterns, but many aspects of the assessment of ridden behavior still rely on subjective judgement. The development of a list of behaviors exhibited by ridden horses, a ridden horse ethogram, will facilitate recording of observable behavioral events. However, without objective evidence of the relevance of these behavioral events, such a resource has limited value

Metal-macrofauna interactions determine microbial community structure and function in copper contaminated sediments

Peer reviewedPublisher PD

Knockout of Syntaxin-4 in 3T3-L1 adipocytes reveals new insight into GLUT4 trafficking and adiponectin secretion

Adipocytes are key to metabolic regulation, exhibiting insulin-stimulated glucose transport that is underpinned by the insulin-stimulated delivery of glucose transporter type 4 (SLC2A4, also known and hereafter referred to as GLUT4)-containing vesicles to the plasma membrane where they dock and fuse, and increase cell surface GLUT4 levels. Adipocytokines, such as adiponectin, are secreted via a similar mechanism. We used genome editing to knock out syntaxin-4, a protein reported to mediate fusion between GLUT4-containing vesicles and the plasma membrane in 3T3-L1 adipocytes. Syntaxin-4 knockout reduced insulin-stimulated glucose transport and adiponectin secretion by ∼50% and reduced GLUT4 levels. Ectopic expression of haemagglutinin (HA)-tagged GLUT4 conjugated to GFP showed that syntaxin-4-knockout cells retain significant GLUT4 translocation capacity, demonstrating that syntaxin-4 is dispensable for insulin-stimulated GLUT4 translocation. Analysis of recycling kinetics revealed only a modest reduction in the exocytic rate of GLUT4 in knockout cells, and little effect on endocytosis. These analyses demonstrate that syntaxin-4 is not always rate limiting for GLUT4 delivery to the cell surface. In sum, we show that syntaxin-4 knockout results in reduced insulin-stimulated glucose transport, depletion of cellular GLUT4 levels and inhibition of adiponectin secretion but has only modest effects on the translocation capacity of the cells