Herein, we report on the production, characterization, and antioxidant power assessment of carotenoids from the haloarchaeon Halorhabdus utahensis. It was grown at 37 °C and 180 rpm agitation in halobacteria medium supplemented with glucose, fructose, and xylose, each at concentrations of 0.2%, 1%, and 2%, and the carotenoid yield and composition were investigated. The microorganism produced the carotenoids under all the conditions tested, and their amount followed the order glucose < xylose < fructose. The highest yield was achieved in 2% fructose growth medium with 550.60 ± 7.91 μg/g dry cell and 2428.15 ± 49.33 μg/L. Separation and identification of the carotenoids were performed by RP-HPLC and HPLC/APCI-ITMSⁿ. Bacterioruberin was the main carotenoid detected and accounted for 60.6%, 56.4%, and 58.9% in 2% glucose, 1% xylose, and 2% fructose extracts, respectively. Several geometric isomers of bacterioruberin were distinguished, and representatives of monoanhydrobacterioruberin, and bisanhydrobacterioruberin were also detected. The assignment to cis-isomers was attempted through analysis of the UV/Vis spectra, intensity of cis peaks, and spectral fine structures. The extracts exhibited superoxide scavenging activity higher than butylhydroxytoluene, ascorbic acid, and Trolox, selected as antioxidant references. The anti-hyaluronidase capacity was investigated, and the 2% fructose extract showed the highest activity reaching 90% enzyme inhibition with 1.5 μg. The overall data confirm that Hrd. utahensis can be regarded as an interesting source of antioxidants that can find applications in the food and cosmetic sectors. Full article

Candida auris, a multidrug-resistant fungal pathogen, significantly threatens global public health. Recent studies have identified melanin production, a key virulence factor in many pathogenic fungi that protects against external threats like reactive oxygen species, in C. auris. However, the melanin regulation mechanism remains elusive. This study explores the role of the Ras/cAMP/PKA signaling pathway in C. auris melanization. It reveals that the catalytic subunits Tpk1 and Tpk2 of protein kinase A (PKA) are essential, whereas Ras1, Gpr1, Gpa2, and Cyr1 are not. Under melanin-promoting conditions, the tpk1Δ tpk2Δ strain formed melanin granules in the supernatant akin to the wild-type strain but failed to adhere them properly to the cell wall. This discrepancy is likely due to a decreased expression of chitin-synthesis-related genes. Our findings also show that Tpk1 primarily drives melanization, with Tpk2 having a lesser impact. To corroborate this, we found that C. auris must deploy Tpk1-dependent melanin deposition as a defensive mechanism against antioxidant exposure. Moreover, we confirmed that deletion mutants of multicopper oxidase and ferroxidase genes, previously assumed to influence C. auris melanization, do not directly contribute to the process. Overall, this study sheds light on the role of PKA in C. auris melanization and enhances our understanding of the pathogenicity mechanisms of this emerging fungal pathogen. Full article

Oxidative stress adaptation strategies are important to cell function and are linked to cardiac, neurodegenerative disease, and cancer. Representatives of the Archaea domain are used as model organisms based on their extreme tolerance to oxidants and close evolutionary relationship with eukaryotes. A study of the halophilic archaeon Haloferax volcanii reveals lysine acetylation to be associated with oxidative stress responses. The strong oxidant hypochlorite: (i) stimulates an increase in lysine acetyltransferase HvPat2 to HvPat1 abundance ratios and (ii) selects for lysine deacetylase sir2 mutants. Here we report the dynamic occupancy of the lysine acetylome of glycerol-grown H. volcanii as it shifts in profile in response to hypochlorite. These findings are revealed by the: (1) quantitative multiplex proteomics of the SILAC-compatible parent and Δsir2 mutant strains and (2) label-free proteomics of H26 ‘wild type’ cells. The results show that lysine acetylation is associated with key biological processes including DNA topology, central metabolism, cobalamin biosynthesis, and translation. Lysine acetylation targets are found conserved across species. Moreover, lysine residues modified by acetylation and ubiquitin-like sampylation are identified suggesting post-translational modification (PTM) crosstalk. Overall, the results of this study expand the current knowledge of lysine acetylation in Archaea, with the long-term goal to provide a balanced evolutionary perspective of PTM systems in living organisms. Full article

Hydrogen peroxide (H₂O₂) is a common effector of defense mechanisms against pathogenic infections. However, bacterial factors involved in H₂O₂ tolerance remain unclear. Here we used transposon-directed insertion-site sequencing (TraDIS), a technique allowing the screening of the whole genome, to identify genes implicated in H₂O₂ tolerance in Escherichia coli. Our TraDIS analysis identified 10 mutants with fitness defect upon H₂O₂ exposure, among which previously H₂O₂-associated genes (oxyR, dps, dksA, rpoS, hfq and polA) and other genes with no known association with H₂O₂ tolerance in E. coli (corA, rbsR, nhaA and gpmA). This is the first description of the impact of gpmA, a gene involved in glycolysis, on the susceptibility of E. coli to H₂O₂. Indeed, confirmatory experiments showed that the deletion of gpmA led to a specific hypersensitivity to H₂O₂ comparable to the deletion of the major H₂O₂ scavenger gene katG. This hypersensitivity was not due to an alteration of catalase function and was independent of the carbon source or the presence of oxygen. Transcription of gpmA was upregulated under H₂O₂ exposure, highlighting its role under oxidative stress. In summary, our TraDIS approach identified gpmA as a member of the oxidative stress defense mechanism in E. coli. Full article

Staphylococcus aureus has to cope with oxidative stress during infections. In this study, S. aureus was found to be resistant to 100 mM H₂O₂ during aerobic growth. While KatA was essential for this high aerobic H₂O₂ resistance, the peroxiredoxin AhpC contributed to detoxification of 0.4 mM H₂O₂ in the absence of KatA. In addition, the peroxiredoxins AhpC, Tpx and Bcp were found to be required for detoxification of cumene hydroperoxide (CHP). The high H₂O₂ tolerance of aerobic S. aureus cells was associated with priming by endogenous H₂O₂ levels, which was supported by an oxidative shift of the bacillithiol redox potential to −291 mV compared to −310 mV in microaerophilic cells. In contrast, S. aureus could be primed by sub-lethal doses of 100 µM H₂O₂ during microaerophilic growth to acquire an improved resistance towards the otherwise lethal triggering stimulus of 10 mM H₂O₂. This microaerophilic priming was dependent on increased KatA activity, whereas aerobic cells showed constitutive high KatA activity. Thus, KatA contributes to the high H₂O₂ resistance of aerobic cells and to microaerophilic H₂O₂ priming in order to survive the subsequent lethal triggering doses of H₂O₂, allowing the adaptation of S. aureus under infections to different oxygen environments. Full article

Global climate change has imposed harsh environmental conditions such as drought. Naturally, the most compatible fungal consortia operate synergistically to enhance plant growth and ecophysiological responses against abiotic strains. Yet, little is known about the interactions between phytohormone-producing endophytic fungal symbionts and plant growth under drought stress. The existing research was rationalized to recognize the role of newly isolated drought-resistant, antioxidant-rich endophytic fungal consortia hosting a xerophytic plant, Carthamus oxycantha L., inoculated to Moringa oleifera L. grown under drought stress of 8% PEG (polyethylene glycol-8000). Under drought stress, the combined inoculation of endophytic strain Microdochium majus (WA), Meyerozyma guilliermondi (TG), and Aspergillus aculeatus (TL3) exhibited a significant improvement in growth attributes such as shoot fresh weight (1.71-fold), shoot length (0.86-fold), root length (0.65-fold), dry weight (2.18-fold), total chlorophyll (0.46-fold), and carotenoids (0.87-fold) in comparison to control (8% PEG). Primary and secondary metabolites were also increased in M. oleifera inoculated with endophytic consortia, under drought stress, such as proteins (1.3-fold), sugars (0.58-fold), lipids (0.41-fold), phenols (0.36-fold), flavonoids (0.52-fold), proline (0.6-fold), indole acetic acid (IAA) (4.5-fold), gibberellic acid (GA) (0.7-fold), salicylic acid (SA) (0.8-fold), ascorbic acid (ASA) (1.85-fold), while abscisic acid (ABA) level was decreased (−0.61-fold) in comparison to the control (8% PEG). Under drought stress, combined inoculation (WA, TG, TL3) also promoted the antioxidant activities of enzymes such as ascorbate peroxidase (APX) (3.5-fold), catalase (CAT) activity (1.7-fold), and increased the total antioxidant capacity (TAC) (0.78-fold) with reduced reactive oxygen species (ROS) such as H₂O₂ production (−0.4-fold), compared to control (8% PEG), and stomatal aperture was larger (3.5-fold) with a lesser decrease (−0.02-fold) in water potential. Moreover, combined inoculation (WA, TG, TL3) up regulated the expression of MolHSF3, MolHSF19, and MolAPX genes in M. oleifera under drought stress, compared to the control (8% PEG), is suggestive of an important regulatory role for drought stress tolerance governed by fungal endophytes. The current research supports the exploitation of the compatible endophytic fungi for establishing the tripartite mutualistic symbiosis in M. oleifera to alleviate the adverse effects of drought stress through strong antioxidant activities. Full article

Haloarchaea are extreme halophilic microorganisms belonging to the domain Archaea, phylum Euryarchaeota, and are producers of interesting antioxidant carotenoid compounds. In this study, four new strains of Haloarcula sp., isolated from saline lakes of the Atacama Desert, are reported and studied by high-resolution mass spectrometry (UHPLC-Q-Orbitrap-MS/MS) for the first time. In addition, determination of the carotenoid pigment profile from the new strains of Haloarcula sp., plus two strains of Halorubrum tebenquichense, and their antioxidant activity by means of several methods is reported. The effect of biomass on cellular viability in skin cell lines was also evaluated by MTT assay. The cholinesterase inhibition capacity of six haloarchaea (Haloarcula sp. ALT-23; Haloarcula sp. TeSe-41; Haloarcula sp. TeSe-51; Haloarcula sp. Te Se-89 and Halorubrum tebenquichense strains TeSe-85 and Te Se-86) is also reported for the first time. AChE inhibition IC₅₀ was 2.96 ± 0.08 μg/mL and BuChE inhibition IC₅₀ was 2.39 ± 0.09 μg/mL for the most active strain, Halorubrum tebenquichense Te Se-85, respectively, which is more active in BuCHe than that of the standard galantamine. Docking calculation showed that carotenoids can exert their inhibitory activity fitting into the enzyme pocket by their halves, in the presence of cholinesterase dimers. Full article