Genome-scale modeling drives 70-fold improvement of intracellular heme production in Saccharomyces cerevisiae.

Heme is an oxygen carrier and a cofactor of both industrial enzymes and food additives. The intracellular level of free heme is low, which limits the synthesis of heme proteins. Therefore, increasing heme synthesis allows an increased production of heme proteins. Using the genome-scale metabolic model (GEM) Yeast8 for the yeast Saccharomyces cerevisiae, we identified fluxes potentially important to heme synthesis. With this model, in silico simulations highlighted 84 gene targets for balancing biomass and increasing heme production. Of those identified, 76 genes were individually deleted or overexpressed in experiments. Empirically, 40 genes individually increased heme production (up to threefold). Heme was increased by modifying target genes, which not only included the genes involved in heme biosynthesis, but also those involved in glycolysis, pyruvate, Fe-S clusters, glycine, and succinyl-coenzyme A (CoA) metabolism. Next, we developed an algorithmic method for predicting an optimal combination of these genes by using the enzyme-constrained extension of the Yeast8 model, ecYeast8. The computationally identified combination for enhanced heme production was evaluated using the heme ligand-binding biosensor (Heme-LBB). The positive targets were combined using CRISPR-Cas9 in the yeast strain (IMX581-HEM15-HEM14-HEM3-Δshm1-HEM2-Δhmx1-FET4-Δgcv2-HEM1-Δgcv1-HEM13), which produces 70-fold-higher levels of intracellular heme.

Host space, not energy or symbiont size, constrains feather mite abundance across passerine bird species.

Comprehending symbiont abundance among host species is a major ecological endeavour, and the metabolic theory of ecology has been proposed to understand what constrains symbiont populations. We parameterized metabolic theory equations to investigate how bird species' body size and the body size of their feather mites relate to mite abundance according to four potential energy (uropygial gland size) and space constraints (wing area, total length of barbs and number of feather barbs). Predictions were compared with the empirical scaling of feather mite abundance across 106 passerine bird species (26,604 individual birds sampled), using phylogenetic modelling and quantile regression. Feather mite abundance was strongly constrained by host space (number of feather barbs) but not by energy. Moreover, feather mite species' body size was unrelated to the body size of their host species. We discuss the implications of our results for our understanding of the bird-feather mite system and for symbiont abundance in general.

Individual differences in spite predict costly third-party punishment.

OBJECTIVE: Spiteful behaviors are those aimed at inflicting harm on another person while also incurring a cost to the self. Although spite sometimes reflects destructive and socially undesirable behaviors including aggression, the current work sought to examine a potentially socially beneficial aspect of spite: engagement in costly punishment for selfish behavior. METHOD: Four studies used a costly third-party punishment task and measured individual differences in spite, narcissism, Machiavellianism, psychopathy, and motivations for engaging in punishment. RESULTS: Trait spite was positively associated with costly punishment of selfish behavior. That association was independent of other dark personality traits (narcissism, Machiavellianism, psychopathy) and was statistically mediated by a desire for retribution. One of the studies also provided evidence that trait spite was associated with costly punishment of even generous behavior; however, rather than a desire for retribution, that association was mediated by a desire to threaten the person being punished. CONCLUSION: Punishing selfishness and other forms of wrongdoing plays an essential role in cooperative group living. The current work provides new insight into the role spiteful motivations might play in this crucial social behavior.

Antimicrobial Activity and Phytochemical Characterization of Baccharis concava Pers., a Native Plant of the Central Chilean Coast.

Few sclerophyllous plants from the central coast of Chile have been systematically studied. This work describes the phytochemical composition and antimicrobial properties of Baccharis concava Pers. (sin. B. macraei), a shrub found in the first line and near the Pacific coast. B. concava has been traditionally used by indigenous inhabitants of today's central Chile for its medicinal properties. Few reports exist regarding the phytochemistry characterization and biological activities of B. concava. A hydroalcoholic extract of B. concava was prepared from leaves and small branches. Qualitative phytochemical characterization indicated the presence of alkaloids, steroids, terpenoids, flavonoids, phenolic, and tannin compounds. The antimicrobial activity of this extract was assessed in a panel of microorganisms including Gram-positive bacteria, Gram-negative bacteria, and pathogenic yeasts. The extract displayed an important antimicrobial effect against Gram-positive bacteria, Candida albicans, and Cryptococcus neoformans but not against Gram-negatives, for which an intact Lipopolysaccharide is apparently the determinant of resistance to B. concava extracts. The hydroalcoholic extract was then fractionated through a Sephadex LH-20/methanol-ethyl acetate column. Afterward, the fractions were pooled according to a similar pattern visualized by TLC/UV analysis. Fractions obtained by this criterion were assessed for their antimicrobial activity against Staphylococcus aureus. The fraction presenting the most antimicrobial activity was HPLC-ESI-MS/MS, obtaining molecules related to caffeoylquinic acid, dicaffeoylquinic acid, and quercetin, among others. In conclusion, the extracts of B. concava showed strong antimicrobial activity, probably due to the presence of metabolites derived from phenolic acids, such as caffeoylquinic acid, and flavonoids, such as quercetin, which in turn could be responsible for helping with wound healing. In addition, the development of antimicrobial therapies based on the molecules found in B. concava could help to combat infection caused by pathogenic yeasts and Gram-positive bacteria, without affecting the Gram-negative microbiota.

Sodium and lithium exert differential effects on the central carbon metabolism of Debaryomyces hansenii through the glyoxylate shunt regulation.

Debaryomyces hansenii is a halotolerant/halophilic yeast usually found in salty environments. The yeast accumulated sodium at high concentrations, which improved growth in salty media. In contrast, lithium was toxic even at low concentrations and its presence prevented cell proliferation. To analyse the responses to both cations, metabolite levels, enzymatic activities and gene expression were determined, showing that NaCl and LiCl trigger different cellular responses. At high concentrations of NaCl (0.5 or 1.5 M) cells accumulated higher amounts of the intermediate metabolites glyoxylate and malate and, at the same time, the levels of intracellular oxoglutarate decreased. Additionally, 0.5 M NaCl increased the activity of the enzymes isocitrate lyase and malate synthase involved in the synthesis of glyoxylate and malate respectively and decreased the activity of isocitrate dehydrogenase. Moreover, transcription of the genes coding for isocitrate lyase and malate synthase was activated by NaCl. Also, cells accumulated phosphate upon NaCl exposure. None of these effects was provoked when LiCl (0.1 or 0.3 M) was used instead of NaCl. Lithium induced accumulation of higher amounts of oxoglutarate and decreased the concentrations of glyoxylate and malate to non-detectable levels. Cells incubated with lithium also showed higher activity of the isocitrate dehydrogenase and neither increased isocitrate lyase and malate synthase activities nor the transcription of the corresponding genes. In summary, we show that sodium, but not lithium, up regulates the shunt of the glyoxylic acid in D. hansenii and we propose that this is an important metabolic adaptation to thrive in salty environments.

Transcriptomic profiling of the yeast Komagataella phaffii in response to environmental alkalinization.

BACKGROUND: Adaptation to alkalinization of the medium in fungi involves an extensive remodeling of gene expression. Komagataella phaffii is an ascomycetous yeast that has become an organism widely used for heterologous protein expression. We explore here the transcriptional impact of moderate alkalinization in this yeast, in search of suitable novel promoters able to drive transcription in response to the pH signal. RESULTS: In spite of a minor effect on growth, shifting the cultures from pH 5.5 to 8.0 or 8.2 provokes significant changes in the mRNA levels of over 700 genes. Functional categories such as arginine and methionine biosynthesis, non-reductive iron uptake and phosphate metabolism are enriched in induced genes, whereas many genes encoding iron-sulfur proteins or members of the respirasome were repressed. We also show that alkalinization is accompanied by oxidative stress and we propose this circumstance as a common trigger of a subset of the observed changes. PHO89, encoding a Na+/Pi cotransporter, appears among the most potently induced genes by high pH. We demonstrate that this response is mainly based on two calcineurin-dependent response elements located in its promoter, thus indicating that alkalinization triggers a calcium-mediated signal in K. phaffii. CONCLUSIONS: This work defines in K. phaffii a subset of genes and diverse cellular pathways that are altered in response to moderate alkalinization of the medium, thus setting the basis for developing novel pH-controlled systems for heterologous protein expression in this fungus.

Improving therapeutic protein secretion in the probiotic yeast Saccharomyces boulardii using a multifactorial engineering approach.

The probiotic yeast Saccharomyces boulardii (Sb) is a promising chassis to deliver therapeutic proteins to the gut due to Sb's innate therapeutic properties, resistance to phage and antibiotics, and high protein secretion capacity. To maintain therapeutic efficacy in the context of challenges such as washout, low rates of diffusion, weak target binding, and/or high rates of proteolysis, it is desirable to engineer Sb strains with enhanced levels of protein secretion. In this work, we explored genetic modifications in both cis- (i.e. to the expression cassette of the secreted protein) and trans- (i.e. to the Sb genome) that enhance Sb's ability to secrete proteins, taking a Clostridioides difficile Toxin A neutralizing peptide (NPA) as our model therapeutic. First, by modulating the copy number of the NPA expression cassette, we found NPA concentrations in the supernatant could be varied by sixfold (76-458 mg/L) in microbioreactor fermentations. In the context of high NPA copy number, we found a previously-developed collection of native and synthetic secretion signals could further tune NPA secretion between 121 and 463 mg/L. Then, guided by prior knowledge of S. cerevisiae's secretion mechanisms, we generated a library of homozygous single gene deletion strains, the most productive of which achieved 2297 mg/L secretory production of NPA. We then expanded on this library by performing combinatorial gene deletions, supplemented by proteomics experiments. We ultimately constructed a quadruple protease-deficient Sb strain that produces 5045 mg/L secretory NPA, an improvement of > tenfold over wild-type Sb. Overall, this work systematically explores a broad collection of engineering strategies to improve protein secretion in Sb and highlights the ability of proteomics to highlight under-explored mediators of this process. In doing so, we created a set of probiotic strains that are capable of delivering a wide range of protein titers and therefore furthers the ability of Sb to deliver therapeutics to the gut and other settings to which it is adapted.

A deeper look at low-frequency contact ultrasonic debridement in the clinical management of patients with diabetic foot ulcers.

The current study aimed to evaluate the dispersal of solution and microbes (aerosol) in the clinical environment during treatment with Low-frequency contact ultrasonic debridement (LFCUD) with or without suction attachment in patients with diabetic foot ulcers (DFUs). We performed 20 treatments in 10 patients divided into two groups to receive the proposed LFCUD modalities. We measured the microbial load of the environment pre-treatment (sample M1), during treatment with each LFCUD modality (sample M2) and post-treatment (sample M3). The use of LFCUD debridement without a suction attachment results in significantly higher immediate contamination of the clinic environment than the suction attachment, particularly during the procedure (1.70 ± 0.98 log 10 CFU/mL versus 0.77 ± 0.85 log 10 CFU/mL, p = 0.035). When suction is not applied, there are statistically significant differences depending on whether the DFUs are neuropathic or neuroischemic, finding a greater number of microorganisms with high loads in neuropathic DFUs. We found a statistically significant positive correlation between wound area (r = 0.450, p = 0.047) and TBI (r = 0.651, p = 0.006) with the bacterial load during the LFCUD. Based on our results, we recommend using the personal protective equipment required to protect staff members and patients during treatment with LFCUD and using a suction attachment where clinically possible to reduce clinic environmental pollution, especially in neuropathic DFUs and those with larger areas.

Magnetocaloric Properties in Rare-Earth-Based Metal-Organic Frameworks: Influence of Magnetic Density and Hydrostatic Pressure.

Magnetic refrigeration based on the magnetocaloric effect (MCE) in metal-organic frameworks (MOF) is regarded as an attractive approach to create more sustainable cooling systems with higher efficiency than traditional ones. Here, we report a study of the MCE in a series of rare-earth-based MOFs. We have considered the selection of the rare-earth cation by investigating materials belonging to the α-rare-earth polymeric framework-4 (α-RPF-4) MOF family, synthesized with different rare-earth cations, and observed that paramagnetic moment and saturation magnetization play an important role in enhancing the magnetic entropy change ΔSM. The effect of structural parameters has also been considered by investigating three classes of metal-organic Gd materials built up from different types of inorganic secondary building units, including clusters (as in Gd-UiO-66), one-dimensional (as in α-RPF-4), and layered (as in Gd-LRH) conformations. Moreover, the analysis of the hydrostatic pressure influence reveals a significant increase in the -ΔSM and relative cooling power (RCP) with values between 4.3 and 16.3 and 121-509 J/kg. Specifically, the RCPmax found was ∼683 J/kg for Gd-UiO-66, which is higher than the one recently observed for Gd2SiO5 (649.5 J/kg). The present study demonstrates that the engineering of metal-organic framework systems based on high Gd densities may favor enhancing of magnetocaloric responses even at low pressures, thus promoting a new design strategy for efficient cooling devices.

The role of childhood unpredictability in adult health.

This research differentiated childhood unpredictability (i.e., perceptions of uncertainty or instability due to turbulent environmental changes) from other related constructs to identify its role in adult health. Study 1 (N = 441) showed that, beyond other childhood adversity variables (poverty and adverse childhood experiences or ACEs) and demographic characteristics, perceptions of unpredictability were associated with greater functional disability and worse health-related quality of life (assessed via the CDC's HRQOL Healthy Days measure and the RAND SF-36). Study 2 (N = 564) replicated those findings in a more racially diverse sample and showed that associations with childhood unpredictability held while also controlling for the Big 5 personality traits. Findings suggest that effects of unpredictability were especially pronounced among Hispanic (in Study 1), and Black/African American and low-income participants (in Study 2). Experiencing childhood environments that are perceived to be uncertain, unstable, or uncontrollable may put children on a path toward poor health outcomes in adulthood. Findings advance theories of child adversity and health and identify childhood unpredictability as a potentially valuable target for intervention.

Primary Cutaneous Follicle Center Lymphoma of the Eyelid in a Young Man.

A 20-year-old male presented with a fast-growing nodule in his right inferior eyelid, no relevant history was obtained. Final histopathologic diagnosis of primary cutaneous follicle center lymphoma (CD20+, CD10+, bcl6+, bcl10+, mum1+, PAX5+, and bcl2-) was determined. The patient had a complete negative systemic work-up, and 3 cycles of consisting of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone chemotherapy were completed. The initial histopathologic diagnosis had been a non-Hodgkin diffuse large B-cell lymphoma which is an infrequent lymphoma type for this location too. To our knowledge, this is the youngest person reported presenting with an eyelid primary cutaneous follicle center lymphoma.

Virulence and Metabolism Crosstalk: Impaired Activity of the Type Three Secretion System (T3SS) in a Pseudomonas aeruginosa Crc-Defective Mutant.

Pseudomonas aeruginosa is a ubiquitous nosocomial opportunistic pathogen that harbors many virulence determinants. Part of P. aeruginosa success colonizing a variety of habitats resides in its metabolic robustness and plasticity, which are the basis of its capability of adaptation to different nutrient sources and ecological conditions, including the infected host. Given this situation, it is conceivable that P. aeruginosa virulence might be, at least in part, under metabolic control, in such a way that virulence determinants are produced just when needed. Indeed, it has been shown that the catabolite repression control protein Crc, which together with the RNA chaperon Hfq regulates the P. aeruginosa utilization of carbon sources at the post-transcriptional level, also regulates, directly or indirectly, virulence-related processes in P. aeruginosa. Among them, Crc regulates P. aeruginosa cytotoxicity, likely by modulating the activity of the Type III Secretion System (T3SS), which directly injects toxins into eukaryotic host cells. The present work shows that the lack of Crc produces a Type III Secretion-defective phenotype in P. aeruginosa. The observed impairment is a consequence of a reduced expression of the genes encoding the T3SS, together with an impaired secretion of the proteins involved. Our results support that the impaired T3SS activity of the crc defective mutant is, at least partly, a consequence of a defective protein export, probably due to a reduced proton motive force. This work provides new information about the complex regulation of the expression and the activity of the T3SS in P. aeruginosa. Our results highlight the need of a robust bacterial metabolism, which is defective in the ∆crc mutant, to elicit complex and energetically costly virulence strategies, as that provided by the T3SS.

Utilizing intramammary Melaleuca alternifolia as an organic internal sealant for dry-off therapy in Murrah buffaloes.

The effects of intramammary dry cow therapy based on the administration of 5% Melaleuca alternifolia tea tree essential oil (TTO) as an internal teat sealant to Murrah cows were evaluated. A longitudinal prospective and retrospective negative control study was performed using 12 buffaloes from a total of 20 Murrah buffaloes on an organic farm, with the cow used as a control for herself. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for treatments with pure oil (TTO) and medication containing 5% TTO (O5) were determined. The buffaloes were clinically examined, and the teats were evaluated using thermography and ultrasound. Udder health was monitored during the first 100 days in milk (DIM) using milk somatic cell count (SCC) and California mastitis test (CMT). Laboratory tests against standard strains Staphylococcus aureus ATCC®25,923™, Escherichia coli ATCC®25,922™, and wild bacterial strains showed maximum MIC values of 50 µL/mL for the TTO and O5 treatments. One wild-type S. aureus strain showed no MBC. No adverse effects were observed after the intramammary application of TTO. The CMT and SCC values were similar (P > 0.05) for all observations. The medication containing 5% TTO was effective in vitro and compatible with the intramammary tissue in vivo of Murrah buffaloes. TTO was safe, not inducing inflammatory processes or other modifications of the teat detectable by thermography or ultrasound. It was able to protect buffaloes during the dry period under field conditions, demonstrating potential use as a teat sealant for organic farms.

Engineering precursor supply for the high-level production of ergothioneine in Saccharomyces cerevisiae.

Ergothioneine (ERG) is an unusual sulfur-containing amino acid. It is a potent antioxidant, which shows great potential for ameliorating neurodegenerative and cardiovascular diseases. L-ergothioneine is rare in nature, with mushrooms being the primary dietary source. The chemical synthesis process is complex and expensive. Alternatively, ERG can be produced by fermentation of recombinant microorganisms engineered for ERG overproduction. Here, we describe the engineering of S. cerevisiae for high-level ergothioneine production on minimal medium with glucose as the only carbon source. To this end, metabolic engineering targets in different layers of the amino acid metabolism were selected based on literature and tested. Out of 28 targets, nine were found to improve ERG production significantly by 10%-51%. These targets were then sequentially implemented to generate an ergothioneine-overproducing yeast strain capable of producing 106.2 ± 2.6 mg/L ERG in small-scale cultivations. Transporter engineering identified that the native Aqr1 transporter was capable of increasing the ERG production in a yeast strain with two copies of the ERG biosynthesis pathway, but not in the strain that was further engineered for improved precursor supply. Medium optimization indicated that additional supplementation of pantothenate improved the strain's productivity further and that no supplementation of amino acid precursors was necessary. Finally, the engineered strain produced 2.39 ± 0.08 g/L ERG in 160 h (productivity of 14.95 ± 0.49 mg/L/h) in a controlled fed-batch fermentation without supplementation of amino acids. This study paves the way for the low-cost fermentation-based production of ergothioneine.

Detailed Structural Features of the Perovskite-Related Halide RbPbI3 for Solar Cell Applications.

All-inorganic lead halide perovskites like CsPbBr3, CsPbI3, or RbPbI3 are good replacements for the classical hybrid organic-inorganic perovskites like CH3NH3PbI3, susceptible to fast degradation in the presence of humid air. They also exhibit outstanding light absorption properties suitable for solar energy applications. Here, we describe the synthesis of RbPbI3 by mechanochemical procedures with green credentials, avoiding toxic or expensive organic solvents; this specimen exhibits excellent crystallinity. We report neutron powder diffraction data, essential to revisit some subtle structural features around room temperature (200-400 K). In all these regimes, the orthorhombic Pnma crystal structure is characterized by the presence along the b direction of the crystal of double rows of edge-sharing PbI6 octahedra. The lone electron pairs of Pb2+ ions have a strong stereochemical effect on the PbI6 octahedral distortion. The relative covalency of Rb-I versus Pb-I bonds shows that the Pb-I-related motions are more rigid than Rb-I-related vibrations, as seen in the Debye temperatures from the evolution of the anisotropic displacements. The optical gap, measured by diffuse reflectance UV-vis spectroscopy, is ∼2.51 eV and agrees well with ab initio calculations. The thermoelectric Seebeck coefficient is 3 orders of magnitude larger than that of other halide perovskites, with a value of ∼117,000 µV·K-1 at 460 K.

Perceptions of childhood unpredictability, delay discounting, risk-taking, and adult externalizing behaviors: A life-history approach.

Guided by principles from life-history theory, theories of adaptive calibration provide an overarching theoretical framework for understanding the developmental roots of impulsivity and externalizing psychopathology. The current research provides evidence for robust associations between perceptions of childhood unpredictability, delay discounting (Studies 1a and 1b), and adult externalizing traits and behaviors (Study 2). Both associations were observed while controlling for perceptions of the harshness of childhood environments, as well as a range of demographic characteristics. The association with externalizing traits and behavior was observed over and above current mood and depressive symptoms. Study 2 also replicated a previously documented association between changes in maternal employment, residence, and cohabitation during childhood and externalizing behavior and, furthermore, suggested that this association was mediated by perceptions of unpredictability. These studies provided no evidence for links between perceived childhood unpredictability and basic forms of risk-taking (Studies 1a and 1c). This research adds to a growing body of work leveraging principles from life-history theory to demonstrate links between childhood experiences, impulsivity, and potentially debilitating forms of mental illness. This work also highlights the value of assessing people's perceptions of their childhood environments.

Large Enhancement of Critical Current in Superconducting Devices by Gate Voltage.

Significant control over the properties of a high-carrier density superconductor via an applied electric field has been considered infeasible due to screening of the field over atomic length scales. Here, we demonstrate an enhancement of up to 30% in critical current in a back-gate tunable NbN micro- and nano superconducting bridges. Our suggested plausible mechanism of this enhancement in critical current based on surface nucleation and pinning of Abrikosov vortices is consistent with expectations and observations for type-II superconductor films with thicknesses comparable to their coherence length. Furthermore, we demonstrate an applied electric field-dependent infinite electroresistance and hysteretic resistance. Our work presents an electric field driven enhancement in the superconducting property in type-II superconductors which is a crucial step toward the understanding of field-effects on the fundamental properties of a superconductor and its exploitation for logic and memory applications in a superconductor-based low-dissipation digital computing paradigm.

Fosfomycin Resistance Evolutionary Pathways of Stenotrophomonas maltophilia in Different Growing Conditions.

The rise of multidrug-resistant Gram-negative pathogens and the lack of novel antibiotics to address this problem has led to the rescue of old antibiotics without a relevant use, such as fosfomycin. Stenotrophomonas maltophilia is a Gram-negative, non-fermenter opportunistic pathogen that presents a characteristic low susceptibility to several antibiotics of common use. Previous work has shown that while the so-far described mechanisms of fosfomycin resistance in most bacteria consist of the inactivation of the target or the transporters of this antibiotic, as well as the production of antibiotic-inactivating enzymes, these mechanisms are not selected in S. maltophilia fosfomycin-resistant mutants. In this microorganism, fosfomycin resistance is caused by the inactivation of enzymes belonging to its central carbon metabolism, hence linking metabolism with antibiotic resistance. Consequently, it is relevant to determine how different growing conditions, including urine and synthetic sputum medium that resemble infection, could impact the evolutionary pathways towards fosfomycin resistance in S. maltophilia. Our results show that S. maltophilia is able to acquire high-level fosfomycin resistance under all tested conditions. However, although some of the genetic changes leading to resistance are common, there are specific mutations that are selected under each of the tested conditions. These results indicate that the pathways of S. maltophilia evolution can vary depending on the infection point and provide information for understanding in more detail the routes of fosfomycin resistance evolution in S. maltophilia.

Debaryomyces hansenii: an old acquaintance for a fresh start in the era of the green biotechnology.

The halophilic yeast Debaryomyces hansenii has been studied for several decades, serving as eukaryotic model for understanding salt and osmotic tolerance. Nevertheless, lack of consensus among different studies is found and, sometimes, contradictory information derived from studies performed in very diverse conditions. These two factors hampered its establishment as the key biotechnological player that was called to be in the past decade. On top of that, very limited (often deficient) engineering tools are available for this yeast. Fortunately Debaryomyces is again gaining momentum and recent advances using highly instrumented lab scale bioreactors, together with advanced -omics and HT-robotics, have revealed a new set of interesting results. Those forecast a very promising future for D. hansenii in the era of the so-called green biotechnology. Moreover, novel genetic tools enabling precise gene editing on this yeast are now available. In this review, we highlight the most recent developments, which include the identification of a novel gene implicated in salt tolerance, a newly proposed survival mechanism for D. hansenii at very high salt and limiting nutrient concentrations, and its utilization as production host in biotechnological processes.

Discovery of inhibitors of Pseudomonas aeruginosa virulence through the search for natural-like compounds with a dual role as inducers and substrates of efflux pumps.

Multidrug efflux pumps are ancient elements encoded in every genome, from bacteria to humans. In bacteria, in addition to antibiotics, efflux pumps extrude a wide range of substrates, including quorum sensing signals, bacterial metabolites, or plant-produced compounds. This indicates that their original functions may differ from their recently acquired role in the extrusion of antibiotics during human infection. Concerning plant-produced compounds, some of them are substrates and inducers of the same efflux pump, suggesting a coordinated plant/bacteria coevolution. Herein we analyse the ability of 1243 compounds from a Natural Product-Like library to induce the expression of P. aeruginosa mexCD-oprJ or mexAB-oprM efflux pumps' encoding genes. We further characterized natural-like compounds that do not trigger antibiotic resistance in P. aeruginosa and that act as virulence inhibitors, choosing those that were not only inducers but substrates of the same efflux pump. Four compounds impair swarming motility, exotoxin secretion through the Type 3 Secretion System (T3SS) and the ability to kill Caenorhabditis elegans, which might be explained by the downregulation of genes encoding flagellum and T3SS. Our results emphasize the possibility of discovering new anti-virulence drugs by screening natural or natural-like libraries for compounds that behave as both, inducers and substrates of efflux pumps.