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Treatment of Staphylococcus aureus infections is complicated by the development of antibiotic tolerance, a consequence of the ability of S. aureus to enter into a nongrowing, dormant state in which the organisms are referred to as persisters. We report that the clinically approved anthelmintic agent bithionol kills methicillin-resistant S. aureus (MRSA) persister cells, which correlates with its ability to disrupt the integrity of Gram-positive bacterial membranes. Critically, bithionol exhibits significant selectivity for bacterial compared with mammalian cell membranes. All-atom molecular dynamics (MD) simulations demonstrate that the selectivity of bithionol for bacterial membranes correlates with its ability to penetrate and embed in bacterial-mimic lipid bilayers, but not in cholesterol-rich mammalian-mimic lipid bilayers. In addition to causing rapid membrane permeabilization, the insertion of bithionol increases membrane fluidity. By using bithionol and nTZDpa (another membrane-active antimicrobial agent), as well as analogs of these compounds, we show that the activity of membrane-active compounds against MRSA persisters positively correlates with their ability to increase membrane fluidity, thereby establishing an accurate biophysical indicator for estimating antipersister potency. Finally, we demonstrate that, in combination with gentamicin, bithionol effectively reduces bacterial burdens in a mouse model of chronic deep-seated MRSA infection. This work highlights the potential repurposing of bithionol as an antipersister therapeutic agent.

The folding and insertion of integral β-barrel membrane proteins into the outer membrane of Gram-negative bacteria is required for viability and bacterial pathogenesis. Unfortunately, the lack of selective and potent modulators to dissect β-barrel folding in vivo has hampered our understanding of this fundamental biological process. Here, we characterize amonoclonal antibody that selectively inhibits an essential component of the Escherichia coli β-barrel assembly machine, BamA. In the absence of complement or other immune factors, the unmodified antibody MAB1 demonstrates bactericidal activity against an E. coli strain with truncated LPS. Direct binding of MAB1 to an extracellular BamA epitope inhibits its β-barrel folding activity, induces periplasmic stress, disrupts outer membrane integrity, and kills bacteria. Notably, resistance to MAB1-mediated killing reveals a link between outermembrane fluidity and protein folding by BamA in vivo, underscoring the utility of this antibody for studying β-barrel membrane protein folding within a living cell. Identification of this BamA antagonist highlights the potential for new mechanisms of antibiotics to inhibit Gram-negative bacterial growth by targeting extracellular epitopes.

Moringa oleifera Lam . is a common edible plant, famous for several nutritional and therapeutic benefits. This study investigates the salt -induced modulations in plant growth, physio-biochemical responses, and antioxidant performance of M. oleifera grown under 0, 50, and 100 mM NaCl concentrations. Results showed that the plant effectively managed moderate salinity (50 mM NaCl) by maintaining succulence, weight ratios, and biomass allocation patterns of both shoot and root with minimal reduction in dry biomass. However, high salinity (100 mM NaCl) remarkably declined all growth parameters. The plant accumulated more Na + and Cl − , while less K + under salinity as compared to the control. Consequently, osmotic potentials of both root and leaf decreased under salinity, which was corroborated by the high amount of proline and soluble sugars. Increased level of H 2 O 2 with significantly unchanged membrane fluidity indicating its role in perceiving and managing stress at moderate salinity. In addition, increased activities of superoxide dismutase, and catalase, with increased glutathione and flavonoid contents suggest an integrated participation of both enzymatic and non-enzymatic antioxidant components in regulating ROS. On the other hand, high salinity caused an outburst of ROS indicated by high H 2 O 2 , MDA, and electrolyte leakage. As a response, moringa drastically increased the activities of all antioxidant enzymes and contents of antioxidant molecules including ascorbic acid, glutathione, total phenols, and flavonoids with high radical scavenging and reducing power capacities. However, a considerable amount of energy was used in such management resulting in a significant growth reduction at 100 mM NaCl. This study suggests that moringa effectively resisted moderate salinity by modulating physio-biochemical attributes and effectively managing ion toxicity and oxidative stress. Salt stress also enhanced the medicinal potentials of moringa by increasing the contents of antioxidant compounds including ascorbic acid, glutathione, total phenols, and flavonoids and their resulting activities. It can be grown on degraded/ saline lands and biomass of this plant can be used for edible and medicinal purposes, besides providing other benefits in a global climate change scenario.

Carotenoids are associated with several important biological functions as antenna pigments in photosynthesis or protectives against oxidative stress. Occasionally they were also discussed as part of the cold adaptation mechanism of bacteria. For two Staphylococcus xylosus strains we demonstrated an increased content of staphyloxanthin and other carotenoids after growth at 10 °C but no detectable carotenoids after grow at 30 °C. By in vivo measurements of generalized polarization and anisotropy with two different probes Laurdan and TMA-DPH we detected a strong increase in membrane order with a simultaneous increase in membrane fluidity at low temperatures accompanied by a broadening of the phase transition. Increased carotenoid concentration was also correlated with an increased resistance of the cells against freeze-thaw stress. In addition, the fatty acid profile showed a moderate adaptation to low temperature by increasing the portion of anteiso-branched fatty acids. The suppression of carotenoid synthesis abolished the effects observed and thus confirmed the causative function of the carotenoids in the modulation of membrane parameters. A differential transcriptome analysis demonstrated the upregulation of genes involved in carotenoid syntheses under low temperature growth conditions. The presented data suggests that upregulated synthesis of carotenoids is a constitutive component in the cold adaptation strategy of Staphylococcus xylosus and combined with modifications of the fatty acid profile constitute the adaptation to grow under low temperature conditions.

Production of lactic acid bacteria starters for manufacturing food, probiotic, and chemical products requires the application of successive steps: fermentation, concentration, stabilization, and storage. Despite process optimization, losses of bacterial viability and functional activities are observed after stabilization and storage steps due to cell exposure to environmental stresses (thermal, osmotic, mechanical, and oxidative). Bacterial membrane is the primary target for injury and its damage is highly dependent on its physical properties and lipid organization. Membrane fluidity is a key property for maintaining cell functionality, and depends on lipid composition and cell environment. Extensive evidence has been reported on changes in membrane fatty acyl chains when modifying fermentation conditions. However, a deep characterization of membrane physical properties and their evolution following production processes is scarcely reported. Therefore, the aims of this mini-review are (i) to define the membrane fluidity and the methods used to assess it and (ii) to summarize the effect of environmental conditions on membrane fluidity and the resulting impact on the resistance of lactic acid bacteria to the stabilization processes. This will make it possible to highlight existing gaps of knowledge and opens up novel approaches for future investigations.

Cholesterol is an integral component of eukaryotic cell membranes and a key molecule in controlling membrane fluidity, organization, and other physicochemical parameters. It also plays a regulatory function in antibiotic drug resistance and the immune response of cells against viruses, by stabilizing the membrane against structural damage. While it iswell understood that, structurally, cholesterol exhibits a densification effect on fluid lipid membranes, its effects on membrane bending rigidity are assumed to be nonuniversal; i.e., cholesterol stiffens saturated lipid membranes, but has no stiffening effect on membranes populated by unsaturated lipids, such as 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC). This observation presents a clear challenge to structure–property relationships and to our understanding of cholesterol-mediated biological functions. Here, using a comprehensive approach—combining neutron spin-echo (NSE) spectroscopy, solid-state deuterium NMR (²H NMR) spectroscopy, and molecular dynamics (MD) simulations—we report that cholesterol locally increases the bending rigidity of DOPC membranes, similar to saturated membranes, by increasing the bilayer’s packing density. All three techniques, inherently sensitive to mesoscale bending fluctuations, show up to a threefold increase in effective bending rigidity with increasing cholesterol content approaching a mole fraction of 50%. Our observations are in good agreement with the known effects of cholesterol on the area-compressibility modulus and membrane structure, reaffirming membrane structure–property relationships. The current findings point to a scale-dependent manifestation of membrane properties, highlighting the need to reassess cholesterol’s role in controlling membrane bending rigidity over mesoscopic length and time scales of important biological functions, such as viral budding and lipid–protein interactions.

The properties of cell membranes are determined mostly by the types of fatty acids that they contain. Bodhicharla et al. report that a key regulator of membrane fluidity, the PAQR-2/IGLR-2 protein complex... Maintenance of membrane properties is an essential aspect of cellular homeostasis of which the regulatory mechanisms remain mostly uncharacterized. In Caenorhabditis elegans, the PAQR-2 and IGLR-2 proteins act together as a plasma membrane sensor that responds to decreased fluidity by promoting fatty acid desaturation, hence restoring membrane fluidity. Here, we used mosaic analysis for paqr-2 and iglr-2, and tissue-specific paqr-2 expression, to show that membrane homeostasis is achieved cell nonautonomously. Specifically, we found that expression of paqr-2 in the hypodermis, gonad sheath cells, or intestine is sufficient to suppress systemic paqr-2 mutant phenotypes, including tail tip morphology, membrane fluidity in intestinal cells, cold and glucose intolerance, vitellogenin transport to the germline, germ cell development, and brood size. Finally, we show that the cell nonautonomous regulation of membrane homeostasis is conserved in human cells: HEK293 cells that express AdipoR2, a homolog of paqr-2, are able to normalize membrane fluidity in distant cells where AdipoR2 has been silenced. Finally, using C. elegans mutants and small interfering RNA against Δ9 stearoyl-CoA desaturase in HEK293 cells, we show that Δ9 desaturases are essential for the cell nonautonomous maintenance of membrane fluidity. We conclude that cells are able to share membrane components even when they are not in direct contact with each other, and that this contributes to the maintenance of membrane homeostasis in C. elegans and human cells.

The only way to visually observe cellular viscosity, which can greatly influence biological reactions and has been linked to several human diseases, is through viscosity imaging. Imaging cellular viscosity has allowed the mapping of viscosity in cells, and the next frontier is targeted viscosity imaging of organelles and their microenvironments. Here we present a fluorescent molecular rotor/FLIM framework to image both organellar viscosity and membrane fluidity, using a combination of chemical targeting and organelle extraction. For demonstration, we image matrix viscosity and membrane fluidity of mitochondria, which have been linked to human diseases, including Alzheimer's Disease and Leigh's syndrome. We find that both are highly dynamic and responsive to small environmental and physiological changes, even under non-pathological conditions. This shows that neither viscosity nor fluidity can be assumed to be fixed and underlines the need for single-cell, and now even single-organelle, imaging.

Freezing lactic acid bacteria often leads to cell death and loss of technological properties. Our objective was to provide an in-depth characterization of the biophysical properties of the Lactobacillus delbrueckii subsp. bulgaricus membrane in relation to its freeze resistance. Freezing was represented as a combination of cold and osmotic stress. This work investigated the relative incidence of increasing sucrose concentrations coupled or not with subzero temperatures without ice nucleation on the biological and biophysical responses of two strains with different membrane fatty acid compositions and freeze resistances. Following exposure of bacterial cells to the highest sucrose concentration, the sensitive strain exhibited a survival rate of less than 10 % and 5 h of acidifying activity loss. Similar biological activity losses were observed upon freeze-thawing and after osmotic treatment for each strain thus highlighting osmotic stress as the main source of cryoinjury. The direct measurement of membrane fluidity by fluorescence anisotropy was linked to membrane lipid organization characterized by FTIR spectroscopy. Both approaches made it possible to investigate the specific contributions of the membrane core and the bilayer external surface to cell degradation caused by cold and osmotic stress. Cold-induced membrane rigidification had no significant implication on bacterial freeze-thaw resistance. Interactions between extracellular sucrose and membrane phospholipid headgroups under osmotic stress were also observed. Such interactions were more evident in the sensitive strain and when increasing sucrose concentration, thus suggesting membrane permeabilization. The relevance of biophysical properties for elucidating mechanisms of cryoinjury and cryoprotection is discussed.

Plants, algae, and photosynthetic bacteria experience frequent changes in environment. The ability to survive depends on their capacity to acclimate to such changes. In particular, fluctuations in temperature affect the fluidity of cytoplasmic and thylakoid membranes. The molecular mechanisms responsible for the perception of changes in membrane fluidity have not been fully characterized. However, the understanding of the functions of the individual genes for fatty acid desaturases in cyanobacteria and plants led to the directed mutagenesis of such genes that altered the membrane fluidity of cytoplasmic and thylakoid membranes. Characterization of the photosynthetic properties of the transformed cyanobacteria and higher plants revealed that lipid unsaturation is essential for protection of the photosynthetic machinery against environmental stresses, such as strong light, salt stress, and high and low temperatures. The unsaturation of fatty acids enhances the repair of the damaged photosystem II complex under stress conditions. In this review, we summarize the knowledge on the mechanisms that regulate membrane fluidity, on putative sensors that perceive changes in membrane fluidity, on genes that are involved in acclimation to new sets of environmental conditions, and on the influence of membrane properties on photosynthetic functions.