Temporal hierarchy and context-dependence of quorum sensing signal in Pseudomonas aeruginosa

The Gram-negative bacterium Pseudomonas aeruginosa can cause infections in a broad range of hosts including plants, invertebrates and mammals and is an important source of nosocomial infections in humans. We were interested in how differences in the bacteria’s nutritional environment impact bacterial communication and virulence factor production. We grew P. aeruginosa in 96 different conditions in BIOLOG Gen III plates and assayed quorum sensing (QS) signaling over the course of growth. We also quantified pyocyanin and biofilm production and the impact of sub-inhibitory exposure to tobramycin. We found that while 3-oxo-C12 homoserine lactone remained the dominant QS signal to be produced, timing of PQS production differed between media types. Further, whether cells grew predominantly as biofilms or planktonic cells was highly context dependent. Our data suggest that understanding the impact of the nutritional environment on the bacterium can lead to valuable insights into the link between bacterial physiology and pathology

Cerrado and Pantanal fruit flours affect gut microbiota composition in healthy and post-COVID-19 individuals: An in vitro pilot fermentation study

Cerrado and Pantanal plants can provide fruits with high nutritional value and antioxidants. This study aims to evaluate four fruit flours (from jatobá pulp, cumbaru almond, bocaiuva pulp and bocaiuva almond) and their effects on the gut microbiota in healthy (HD) and post-COVID-19 individuals (PC). An in vitro batch system was carried out, the microbiota was analysed by 16S rRNA amplicon sequencing and the short-chain fatty acids (SCFAs) ratio was determined. Furthermore, the effect of jatobá pulp flour oil (JAO) on cell viability, oxidative stress and DNA damage was investigated in a myelomonocytic cell line. Beyond confirming a microbiota imbalance in PC, we identified flourspecific effects: i) reduction of Veillonellaceae with jatobá extract in PC samples;decrease in Akkermansia with jatoba and cumbaru flours; iii) decreasing trend of Faecalibacterium and Ruminococcus with all flours tested, with the exception of the bocaiuva almond in HD samples for Ruminococcus; and iv) increase in Lactobacillus and Bifidobacterium in PC samples with bocaiuva almond flour. JAO displayed antioxidant properties protecting cells from daunorubicin-induced cytotoxicity, oxidative stress, and DNA damage. The promising microbiota modulating abilities of some flours and the chemopreventive effects of JAO deserve to be further explored in human intervention studies

Antibiotic Pollution in Surface Fresh Waters: Occurrence and Effects

Worldwide, antibiotic usage exceeds 100,000 tons per year and there is increasing concern over the fate of these substances. Antibiotics are ubiquitous in the environment and significant concentrations have been detected in fresh waters. In this review, we highlight important aspects of antibiotic pollution in fresh waters: that concentrations of antibiotics in the environment are substantial, that micro-organisms are susceptible to this, that bacteria can evolve resistance in the environment, and that antibiotic pollution affects natural food webs while interacting with other stressors; which taken together poses a number of challenges for environmental scientists. In the literature, we found examples of considerable antibiotic pollution in fresh waters. In the Americas, antibiotic concentrations of up to 15 μg/L have been measured; with higher concentrations reported from European and African studies (over 10 μg/L and 50 μg/L respectively), and in Asian-pacific countries concentrations over 450 μg/L have been detected. While these concentrations might not be deemed harmful to humans, non-target freshwater organisms could be affected by them. Bioassays show that some of the antibiotics found in surface waters affect microbes at concentrations below 10 μg/L. Among the most potent antibiotics are those that prevail in streams and rivers in these concentrations, such as ciprofloxacin. Sub-lethal concentrations might not kill prokaryotes but contribute to increased bacterial resistance and change the composition of single-celled communities, as demonstrated in laboratory experiments. This has implications for the microbial food web (e.g. interactions among and between bacteria and their protozoan consumers) and by extension, larger organisms and ecosystem health. The fact that the effects of antibiotics are extremely context-dependent represents a challenge, particularly for in vitro research. We suggest future research avenues, taking into account food web experiments, antibiotics interacting with one another (and other stressors) and discuss how these can help to answer multi-layered research questions

Intensity interrogation near cutoff resonance for label-free cellular profiling

We report a method enabling intensity-based readout for label-free cellular assays, and realize a reader device with the same footprint as a microtiter plate. For unambiguous resonance intensity measurements in resonance waveguide grating (RWG) sensors, we propose to apply resonances near the substrate cutoff wavelength. This method was validated in bulk refractive index, surface bilayer and G protein-coupled receptor (GPCR) experiments. The significantly reduced size of the reader device opens new opportunities for easy integration into incubators or liquid handling systems

NMR Spectroscopy of Cell Culture, Tissues, and Other Biofluids

NMR spectroscopy can provide a wealth of information on cellular metabolism and is frequently used in metabolomics application that use cultured cells, tissues, and whole organisms. Central to these analyses are the protocols for sample harvest, which incorporate procedures for quenching metabolic processes to preserve samples in a state that is representative of their source. In this chapter, the main considerations are discussed with reference to literature exemplars. In the latter half of the chapter, less commonly studied biofluids that also have specific sample preparation requirements are discussed, with a focus on cerebrospinal fluid, faeces, bile, seminal fluid, and milk.</jats:p

GlnK Facilitates the Dynamic Regulation of Bacterial Nitrogen Assimilation

Ammonium assimilation in Escherichia coli is regulated by two paralogous proteins (GlnB and GlnK), which orchestrate interactions with regulators of gene expression, transport proteins, and metabolic pathways. Yet how they conjointly modulate the activity of glutamine synthetase, the key enzyme for nitrogen assimilation, is poorly understood. We combine experiments and theory to study the dynamic roles of GlnB and GlnK during nitrogen starvation and upshift. We measure time-resolved in vivo concentrations of metabolites, total and posttranslationally modified proteins, and develop a concise biochemical model of GlnB and GlnK that incorporates competition for active and allosteric sites, as well as functional sequestration of GlnK. The model predicts the responses of glutamine synthetase, GlnB, and GlnK under time-varying external ammonium level in the wild-type and two genetic knock-outs. Our results show that GlnK is tightly regulated under nitrogen-rich conditions, yet it is expressed during ammonium run-out and starvation. This suggests a role for GlnK as a buffer of nitrogen shock after starvation, and provides a further functional link between nitrogen and carbon metabolisms

Involvement of the extracellular matrix and integrin signalling proteins in skeletal muscle glucose uptake

Whole-body euglycaemia is partly maintained by two cellular processes that encourage glucose uptake in skeletal muscle; 1) the insulin- and 2) contraction-stimulated pathways, with research suggesting convergence between these two previously separate processes. The normal structural integrity of the skeletal muscle requires an intact actin cytoskeleton as well as integrin-associated proteins, thus those structures are likely fundamental for effective glucose uptake in skeletal muscle. In contrast, excessive extracellular matrix (ECM) remodelling and integrin expression in skeletal muscle may contribute to insulin resistance owing to an increased physical barrier causing reduced nutrient and hormonal flux. This review paper explores the role of the ECM and the actin cytoskeleton in insulin- and contraction-mediated glucose uptake in skeletal muscle. This is a clinically important area of research given that defects in the structural integrity of the ECM and integrin-associated proteins may contribute to loss of muscle function and decreased glucose uptake in type 2 diabetes

Involvement of the extracellular matrix and integrin signalling proteins in skeletal muscle glucose uptake

Whole‐body euglycaemia is partly maintained by two cellular processes that encourage glucose uptake in skeletal muscle, the insulin‐ and contraction‐stimulated pathways, with research suggesting convergence between these two processes. The normal structural integrity of the skeletal muscle requires an intact actin cytoskeleton as well as integrin‐associated proteins, and thus those structures are likely fundamental for effective glucose uptake in skeletal muscle. In contrast, excessive extracellular matrix (ECM) remodelling and integrin expression in skeletal muscle may contribute to insulin resistance owing to an increased physical barrier causing reduced nutrient and hormonal flux. This review explores the role of the ECM and the actin cytoskeleton in insulin‐ and contraction‐mediated glucose uptake in skeletal muscle. This is a clinically important area of research given that defects in the structural integrity of the ECM and integrin‐associated proteins may contribute to loss of muscle function and decreased glucose uptake in type 2 diabetes. [Image: see text

Intracellular Staphylococcus aureus modulates host central carbon metabolism to activate autophagy

Staphylococcus aureus is a facultative intracellular pathogen that invades and replicates within many types of phagocytic and nonphagocytic cells. During intracellular infection, S. aureus is capable of subverting xenophagy and escaping to the cytosol of the host cell. Furthermore, drug-induced autophagy facilitates the intracellular replication of S. aureus, but the reasons behind this are unclear. Here, we have studied the host central carbon metabolism during S. aureus intracellular infection. We found extensive metabolic rerouting and detected several distinct metabolic changes that suggested starvation-induced autophagic flux in infected cells. These changes included increased uptake but lower intracellular levels of glucose and low abundance of several essential amino acids, as well as markedly upregulated glutaminolysis. Furthermore, we show that AMP-activated protein kinase (AMPK) and extracellular signal-regulated kinase (ERK) phosphorylation levels are significantly increased in infected cells. Interestingly, while autophagy was activated in response to S. aureus invasion, most of the autophagosomes detected in infected cells did not contain bacteria, suggesting that S. aureus induces the autophagic flux during cell invasion for energy generation and nutrient scavenging. Accordingly, AMPK inhibition halted S. aureus intracellular proliferation

The change of psychosocial stress factors in families with infants and toddlers during the COVID-19 pandemic. A longitudinal perspective on the CoronabaBY study from Germany

BackgroundOver nearly three years, the COVID-19 pandemic has had a lasting impact on people's lives and mental health worldwide with its far-reaching restrictions and concerns about infections and other personal consequences. Families were particularly affected and showed increased stress and psychological problems. Long-term effects cannot be ruled out. So far, data on young families are sparse. The present longitudinal analysis (n = 932) of the CoronabaBY study investigated the development of parenting stress, parental affective symptoms, and child's mental health in young families with children aged 0–3 years in Germany as well as potential influencing factors.MethodsThe observational study includes two measurement points over the course of the pandemic (baseline and follow-up). Data was collected by app using standardized questionnaires.ResultsN = 932 participants, mainly mothers (94.7%) born in Germany (93.1%) with higher education (61.3% with at least high school diploma) and a comfortable financial situation participated in the longitudinal study. Children were on average 14.7 months old at baseline (SD: 12, range: 1–39 months). While the proportion of parents who perceived the pandemic as stressful decreased significantly from baseline (60%) to follow-up (52.3%), the proportion with parenting stress increased significantly (from 40.1% to 45.4%). Both parental and child mental health problems remained constant over time, with infants crying/feeding/sleeping problems ranging above pre-pandemic comparative data. Most predictive for high parenting stress at follow-up was high parenting stress at baseline. This was also true for parental affective symptoms (depression/anxiety) and child mental health problems.ConclusionsDespite faded pandemic restrictions, parents remained burdened. Support services do not appear to have been sufficient to help families out of their stressful situation. Our results indicate a need for action regarding low-threshold services that effectively reach affected families.Trial registrationThe study was pre-registered in OSF (https://osf.io/search/?q=tksh5&amp;page=1)