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Efflux pump inhibitors are a potential therapeutic strategy for managing antimicrobial resistance and biofilm formation. This article evaluated the effect of carbonyl cyanide m-chlorophenyl hydrazone (CCCP) on the biofilm growth dynamics and the production of virulence factors by Burkholderia pseudomallei . The effects of CCCP on planktonic, growing, and mature biofilm, interaction with antibacterial drugs, and protease and siderophore production were assessed. CCCP MICs ranged between 128 and 256 µM. The CCCP (128 µM) had a synergic effect with all the antibiotics tested against biofilms. Additionally, CCCP reduced ( p  < .05) the biomass of biofilm growth and mature biofilms at 128 and 512 µM, respectively. CCCP also decreased ( p  < .05) protease production by growing (128 µM) and induced ( p  < .05) siderophore release by planktonic cells (128 µM) growing biofilms (12.8 and 128 µM) and mature biofilms (512 µM). CCCP demonstrates potential as a therapeutic adjuvant for disassembling B. pseudomallei biofilms and enhancing drug penetration.

Fungi pose a global threat to humankind due to the increasing emergence of multi-drug-resistant fungi. There is a rising incidence of invasive fungal infections. Due to the structural complexity of fungal cell membranes, only a few classes of antifungal agents are effective and have been approved by the U.S. FDA. Hence, researchers globally are focusing on developing novel strategies to cure fungal infections. One of the potential strategies is the "Trojan horse" approach, which uses the siderophore-mediated iron acquisition (SIA) system to scavenge iron to deliver potent antifungal agents for therapeutics and diagnostics. These siderophore conjugates chelate to iron and are taken up through siderophore-iron transporters, which are overexpressed exclusively on microbes such as bacteria or fungi, but not mammalian cells. Our comprehensive review delves into recent advancements in the design of siderophore-conjugated antifungal agents to gain fungal cell entry. Notably, our focus extends to unraveling the intricate relationship between the structure of natural siderophores or siderophore-like molecules and the resulting antifungal activity. By exploring these design strategies, we aim to contribute to the ongoing discourse on combating drug-resistant fungal infections and advancing the landscape of antifungal theranostics.

In this Review, we explore natural product antibiotics that do more than simply inhibit an active site of an essential enzyme. We review these compounds to provide inspiration for the design of much-needed new antibacterial agents, and examine the complex mechanisms that have evolved to effectively target bacteria, including covalent binders, inhibitors of resistance, compounds that utilize self-promoted entry, those that evade resistance, prodrugs, target corrupters, inhibitors of 'undruggable' targets, compounds that form supramolecular complexes, and selective membrane-acting agents. These are exemplified by β-lactams that bind covalently to inhibit transpeptidases and β-lactamases, siderophore chimeras that hijack import mechanisms to smuggle antibiotics into the cell, compounds that are activated by bacterial enzymes to produce reactive molecules, and antibiotics such as aminoglycosides that corrupt, rather than merely inhibit, their targets. Some of these mechanisms are highly sophisticated, such as the preformed β-strands of darobactins that target the undruggable β-barrel chaperone BamA, or teixobactin, which binds to a precursor of peptidoglycan and then forms a supramolecular structure that damages the membrane, impeding the emergence of resistance. Many of the compounds exhibit more than one notable feature, such as resistance evasion and target corruption. Understanding the surprising complexity of the best antimicrobial compounds provides a roadmap for developing novel compounds to address the antimicrobial resistance crisis by mining for new natural products and inspiring us to design similarly sophisticated antibiotics., (© 2024. Springer Nature Limited.)

Mycoplasma genitalium is a sexually transmitted bacterium associated with urogenital disease syndromes in the US and worldwide. The global rise in drug resistance in M. genitalium necessitates the development of novel drugs to treat this pathogen. To address this need, we have screened extracts from a library of fungal isolates assembled through the University of Oklahoma Citizen Science Soil Collection Program. Analysis of one of the bioactive extracts using bioassay-guided fractionation led to the purification of the compound PF1140 ( 1 ) along with a new and several other known pyridones. The N -hydroxy pyridones are generally regarded as siderophores with high binding affinity for iron(III) under physiological conditions. Results from UV-vis absorption spectroscopy-based titration experiments revealed that 1 complexes with Fe 3+ . As M. genitalium does not utilize iron, we propose that the PF1140-iron complex induces cytotoxicity by facilitating the cellular uptake of iron, which reacts with endogenous hydrogen peroxide to produce toxic hydroxyl radicals.

We designed a tris-catecholate-based siderophore mimic, H 6 -T-CATL, to selectively chelate iron(III) from mitochondrial cytochromes and other iron-containing proteins within cellular matrices. This strategic sequestration aims to trigger apoptosis or ferroptosis in cancer cells through the glutathione (GSH)-dependent release of reduced iron and subsequent ROS-mediated cytotoxicity. Synthesis of H 6 -T-CATL involved precise peptide coupling reactions. Using the Fe(III)-porphyrin model (Fe-TPP-Cl), akin to cytochrome c , we studied H 6 -T-CATL's ability to extract iron(III), yielding a binding constant ( K rel ) of 10 14 for the resulting iron(III) complex (FeIII-T-CATL)3-. This complex readily underwent GSH-mediated reduction to release bioavailable iron(II), which catalyzed Fenton-like reactions generating hydroxyl radicals (˙OH), confirmed by spectroscopic analyses. Our research underscores the potential of H 6 -T-CATL to induce cancer cell death by depleting iron(III) from cellular metalloproteins, releasing pro-apoptotic iron(II). Evaluation across various cancer types, including normal cells, demonstrated H 6 -T-CATL's cytotoxicity through ROS production, mitochondrial dysfunction, and activation of ferroptosis and DNA damage pathways. These findings propose a novel mechanism for cancer therapy, leveraging endogenous iron stores within cells. H 6 -T-CATL emerges as a promising next-generation anticancer agent, exploiting iron metabolism vulnerabilities to induce selective cancer cell death through ferroptosis induction.

Siderophores are structurally unique medicinal natural products and exhibit considerable therapeutic potential. Herein, we report the design and synthesis of azotochelin, a natural siderophore, and an extensive library of azotochelin analogs and their anticancer properties. We modified the carboxylic acid and the aromatic ring of azotochelin using various chemical motifs. We evaluated the cytotoxicity of the compounds against six different cancer cell lines (KB-3-1, SNB-19, MCF-7, K-562, SW-620, and NCI-H460) and a non-cancerous cell line (HEK-293). Among the twenty compounds tested, the IC 50 values of nine compounds (14, 32, 35-40, and 54) were between 0.7 and 2.0 μM against a lung cancer cell line (NCI-H460). Moreover, several compounds showed good cytotoxicity profile (IC 50 <10 μM) against the tested cancer cell lines. The flow cytometry analysis showed that compounds 36 and 38 induced apoptosis in NCI-H460 in a dose-dependent manner. The cell cycle analysis indicated that compounds 36 and 38 significantly arrested the cell cycle at the S phase to block cancer cell proliferation in the NCI-H460 cell line. The study has produced various novel azotochelin analogs that are potentially effective anticancer agents and lead compounds for further synthetic and medicinal chemistry exploration., (© 2024 Wiley-VCH GmbH.)

Many microbes produce siderophores, which are extremely potent weapons capable of stealing iron ions from human tissues, fluids and cells and transferring them into bacteria through their appropriate porins. We have recently designed a multi-block molecule, each block having a dedicated role. The first component is an antimicrobial peptide, whose good effectiveness against some bacterial strains was gradually improved through interactive sequence modifications. Connected to this block is a flexible bio-band, also optimized in length, which terminates in a hydroxyamide unit, a strong metal binder. Thus, the whole molecule brings together two pieces that work synergistically to fight infection. To understand if the peptide unit, although modified with a long tail, preserves the structure and therefore the antimicrobial activity, and to characterize the mechanism of interaction with bio-membrane models mimicking Gram-negative membranes, we performed a set of fluorescence-based experiments and circular dichroism studies, which further supported our design of a combination of two different entities working synergistically. The chelating activity and iron(III) binding of the peptide was confirmed by iron(III) paramagnetic NMR analyses, and through a competitive assay with ethylenediamine-tetra acetic acid by ultraviolet-visible spectroscopy. The complexation parameters, the Michaelis constant K, and the number of sites n, evaluated with spectrophotometric techniques are confirmed by Fe(III) paramagnetic NMR analyses here reported. In conclusion, we showed that the coupling of antimicrobial capabilities with iron-trapping capabilities works well in the treatment of infectious diseases caused by Gram-negative pathogens., Competing Interests: Declaration of competing interest There are no conflicts to declare., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Biofilms commonly develop in immunocompromised patients, which leads to persistent infections that are difficult to treat. In the biofilm state, bacteria are protected against both antibiotics and the host's immune system; currently, there are no therapeutics that target biofilms. In this study, we screened a chemical fraction library representing the natural product capacity of the microbiota of marine egg masses, namely, the moon snail egg collars. This led to the identification of active fractions targeting both Pseudomonas aeruginosa and Staphylococcus aureus biofilms. Subsequent analysis revealed that a subset of these fractions were capable of eradicating preformed biofilms, all against S. aureus . Bioassay-guided isolation led us to identify pseudochelin A, a known siderophore, as a S. aureus biofilm inhibitor with an IC 50 of 88.5 μM. Mass spectrometry-based metabolomic analyses revealed widespread production of pseudochelin A among fractions possessing S. aureus antibiofilm properties. In addition, a key biosynthetic gene involved in producing pseudochelin A was detected on 30% of the moon snail egg collars and pseudochelin A is capable of inhibiting the formation of biofilms (IC 50 50.6 μM) produced by ecologically relevant bacterial strains. We propose that pseudochelin A may have a role in shaping the microbiome or protecting the egg collars from microbiofouling.

The design of siderophore-antibiotic conjugates is a promising strategy to overcome drug resistance in negative bacteria. However, accumulating studies have shown that only those antibiotics acting on the cell wall or cell membrane multiply their antibacterial effects when coupled with siderophores, while antibiotics acting on targets in the cytoplasm of bacteria do not show an obvious enhancement of their antibacterial effects when coupled with siderophores. To explore the causes of this phenomenon, we synthesized several conjugate probes using 3-hydroxypyridin-4(1 H )-ones as siderophores and replacing the antibiotic cargo with 5-carboxyfluorescein (5-FAM) or malachite green (MG) cargo. By monitoring changes in the fluorescence intensity of FAM conjugate 20 in bacteria, the translocation of the conjugate across the outer membranes of Gram-negative pathogens was confirmed. Further, the use of the fluorogen activating protein(FAP)/MG system revealed that 3-hydroxypyridin-4(1 H )-one-MG conjugate 26 was ultimately distributed mainly in the periplasm rather than being translocated into the cytosol of Escherichia coli and Pseudomonas aeruginosa PAO1. Additional mechanistic studies suggested that the uptake of the conjugate involved the siderophore-dependent iron transport pathway and the 3-hydroxypyridin-4(1 H )-ones siderophore receptor-dependent mechanism. Meanwhile, we demonstrated that the conjugation of 3-hydroxypyridin-4(1 H )-ones to the fluorescein 5-FAM can reduce the possibility of the conjugates crossing the membrane layers of mammalian Vero cells by passive diffusion, and the advantages of the mono-3-hydroxypyridin-4(1 H )-ones as a delivery vehicle in the design of conjugates compared to the tri-3-hydroxypyridin-4(1 H )-ones. Overall, this work reveals the localization rules of 3-hydroxypyridin-4(1 H )-ones as siderophores to deliver the cargo into Gram-negative bacteria. It provides a theoretical basis for the subsequent design of siderophore-antibiotic conjugates, especially based on 3-hydroxypyridin-4(1 H )-ones as siderophores.

Multidrug-resistant Acinetobacter baumannii (MDRAB) is an important pathogen that causes nosocomial infections and is resistant to almost all antibiotics, including carbapenems. Cefiderocol is a novel siderophore cephalosporin active against a broad spectrum of gram-negative bacteria. However, the susceptibility of MDRAB to cefiderocol has not yet been reported in Japan. In this study, we measured the minimum inhibitory concentrations (MICs) of antibiotics including cefiderocol against MDRAB clinical isolates collected during a nosocomial outbreak between 2009 and 2010 at the Teikyo University Hospital in Japan. We found that all 10 MDRAB clinical isolates tested were susceptible to cefiderocol, with an MIC range of 0.12 to 1 μg/mL. All the isolates also exhibited resistance to ampicillin-sulbactam and an intermediate resistance to colistin, whereas nine of them were susceptible to tigecycline. DNA sequencing revealed that all strains harbored an OXA-51-like carbapenemase, a major cause of carbapenem resistance in A. baumannii in Japan. In conclusion, this study showed that the cefiderocol susceptibility of MDRAB clinical isolates in Japan was equivalent to that to colistin or tigecycline, and thus cefiderocol is a potential treatment option for MDRAB infections.

Clavatols exhibit a wide range of biological activities due to their diverse structures. A genome mining strategy identified an A5cla cluster from Penicillium sp. MYA5, derived from the Arctic plant Dryas octopetala , is responsible for clavatol biosynthesis. Seven clavatols, including one new clavatol derivate named penicophenone F ( 1 ) and six known clavatols ( 2 - 7 ), were isolated from Penicillium sp. MYA5 using a transcriptome mining strategy. These structures were elucidated by comprehensive spectroscopic analysis. Antibacterial, aldose reductase inhibition, and siderophore-producing ability assays were conducted on compounds 1 - 7 . Compounds 1 and 2 demonstrated inhibitory effects on the ALR2 enzyme with inhibition rates of 75.3% and 71.6% at a concentration of 10 μM, respectively. Compound 6 exhibited antibacterial activity against Staphylococcus aureus and Escherichia coli with MIC values of 4.0 μg/mL and 4.0 μg/mL, respectively. Additionally, compounds 1 , 5 , and 6 also showed potential iron-binding ability.

Antibacterial resistance is a major threat for human health. There is a need for new antibacterials to stay ahead of constantly-evolving resistant bacteria. Nucleic acid therapeutics hold promise as powerful antibiotics, but issues with their delivery hamper their applicability. Here, we exploit the siderophore-mediated iron uptake pathway to efficiently transport antisense oligomers into bacteria. We appended a synthetic siderophore to antisense oligomers targeting the essential acpP gene in Escherichia coli. Siderophore-conjugated PNA and PMO antisense oligomers displayed potent antibacterial properties. Conjugates bearing a minimal siderophore consisting of a mono-catechol group showed equally effective. Targeting the lacZ transcript resulted in dose-dependent decreased β-galactosidase production, demonstrating selective protein downregulation. Applying this concept to Acinetobacter baumannii also showed concentration-dependent growth inhibition. Whole-genome sequencing of resistant mutants and competition experiments with the endogenous siderophore verified selective uptake through the siderophore-mediated iron uptake pathway. Lastly, no toxicity towards mammalian cells was found. Collectively, we demonstrate for the first time that large nucleic acid therapeutics can be efficiently transported into bacteria using synthetic siderophore mimics., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Background: Iron is a vital element for the growth of bacteria. Bacteria use several strategies to scavenge iron, such as siderophores, which are thought to be important virulence components. The mammalian host uses various iron-binding substances to make iron unavailable for bacterial uptake. Deferoxamine (DFO) is a semi-synthetic iron chelator that has been licensed for medical use. Iron chelators like DFO may provide an alternative therapeutic technique for treating Gram-negative bacteria infections, which frequently display multidrug resistance., Objectives: We assumed that iron deprivation or interactions with the cell membrane caused by DFO or increased siderophore synthesis may cause the inhibition or inactivation of proteins and enzymes necessary for critical processes in bacteria. Additionally, we proposed that these bacterial alterations might be the origin of synergistic interactions between DFO and several antibiotics., Material and Methods: To test this hypothesis, we used disc diffusion, broth microdilution and checkerboard synergy testing methods on combinations of DFO with ceftriaxone, cefepime, meropenem, amikacin, levofloxacin, and tigecycline, respectively, in a total of 55 isolates (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Proteus mirabilis strains - 11 isolates for each genus)., Results: No synergistic or antagonistic interactions were observed between DFO and the tested antibiotics in the E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii isolates, while the addition of DFO significantly increased the inhibition zone diameters of cefepime, amikacin, meropenem, tigecycline, and levofloxacin in P. mirabilis isolates. According to the checkerboard synergy results, a synergistic interaction was found between DFO and tigecycline, cefepime and amikacin for P. mirabilis isolates., Conclusions: Among the investigated bacteria, a synergy between antibiotics and DFO was only discovered against P. mirabilis. We do not believe that this entirely disproves our hypothesis, though. The production of siderophores triggered by the increased metabolic activity of actively proliferating bacteria at the infection site may provide better results. Therefore, expanding these investigations and developing infection models through animal testing would be advantageous.

Covering: 2005 to August, 2023Polyamine-containing natural products (NPs) have been isolated from a wide range of terrestrial and marine organisms and most of them exhibit remarkable and diverse activities, including antimicrobial, antiprotozoal, antiangiogenic, antitumor, antiviral, iron-chelating, anti-depressive, anti-inflammatory, insecticidal, antiobesity, and antioxidant properties. Their extraordinary activities and potential applications in human health and agriculture attract increasing numbers of studies on polyamine-containing NPs. In this review, we summarized the source, structure, classification, bioactivities and biosynthesis of polyamine-containing NPs, focusing on the biosynthetic mechanism of polyamine itself and representative polyamine alkaloids, polyamine-containing siderophores with catechol/hydroxamate/hydroxycarboxylate groups, nonribosomal peptide-(polyketide)-polyamine (NRP-(PK)-PA), and NRP-PK-long chain poly-fatty amine (lcPFAN) hybrid molecules.

The growing threat of bacterial infections coupled with the dwindling arsenal of effective antibiotics has heightened the urgency for innovative strategies to combat bacterial pathogens, particularly Gram-negative strains, which pose a significant challenge due to their outer membrane permeability barrier. In this study, we repurpose clinically approved anticancer agents as targeted antibacterials. We report two new siderophore-platinum(IV) conjugates, both of which consist of an oxaliplatin-based Pt(IV) prodrug (oxPt(IV)) conjugated to enterobactin (Ent), a triscatecholate siderophore employed by Enterobacteriaceae for iron acquisition. We demonstrate that l/d-Ent-oxPt(IV) (l/d-EOP) are selectively delivered into the Escherichia col i cytoplasm, achieving targeted antibacterial activity, causing filamentous morphology, and leading to enhanced Pt uptake by bacterial cells but reduced Pt uptake by human cells. d-EOP exhibits enhanced potency compared to oxaliplatin and l-EOP, primarily attributed to the intrinsic antibacterial activity of its non-native siderophore moiety. To further elucidate the antibacterial activity of Ent-Pt(IV) conjugates, we probed DNA damage caused by l/d-EOP and the previously reported cisplatin-based conjugates l/d-Ent-Pt(IV) (l/d-EP). A comparative analysis of these four conjugates reveals a correlation between antibacterial activity and the ability to induce DNA damage. This work expands the scope of Pt cargos targeted to the cytoplasm of Gram-negative bacteria via Ent conjugation, provides insight into the cellular consequences of Ent-Pt(IV) conjugates in E. coli , and furthers our understanding of the potential of Pt-based therapeutics for antibacterial applications.

The low permeability of the outer membrane of Gram-negative bacteria is a serious obstacle to the development of new antibiotics against them. Conjugation of antibiotic with siderophore based on the "Trojan horse strategy" is a promising strategy to overcome the outer membrane obstacle. In this study, series of antibacterial agents were designed and synthesized by conjugating the 3-hydroxypyridin-4(1H)-one based siderophores with cajaninstilbene acid (CSA) derivative 4 which shows good activity against Gram-positive bacteria by targeting their cell membranes but is ineffective against Gram-negative bacteria. Compared to the inactive parent compound 4, the conjugates 45c or 45d exhibits significant improvement in activity against Gram-negative bacteria, including Escherichia coli, Klebsiella pneumoniae and especially P. aeruginosa (minimum inhibitory concentrations, MICs = 7.8-31.25 μM). The antibacterial activity of the conjugates is attributed to the CSA derivative moiety, and the action mechanism is by disruption of bacterial cell membranes. Further studies on the uptake mechanisms showed that the bacterial siderophore-dependent iron transport system was involved in the uptake of the conjugates. In addition, the conjugates 45c and 45d showed a lower cytotoxic effects in vivo and in vitro and a positive therapeutic effect in the treatment of C. elegans infected by P. aeruginosa. Overall, our work describes a new class and a promising 3-hydroxypyridin-4(1H)-one-CSA derivative conjugates for further development as antibacterial agents against Gram-negative bacteria., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Developing new antibiotics and delivery strategies is of critical importance for treating infections caused by Gram-negative bacterial pathogens. Hijacking bacterial iron uptake machinery, such as that of the siderophore enterobactin (Ent), represents one promising approach toward these goals. Here, we report a novel Ent-inspired siderophore-antibiotic conjugate (SAC) employing an alternative siderophore moiety as the delivery vector and demonstrate the potency of our SACs harboring the β-lactam antibiotic ampicillin (Amp) against multiple pathogenic Gram-negative bacterial strains. We establish the ability of N,N',N'' -(nitrilotris(ethane-2,1-diyl))tris(2,3-dihydroxybenzamide) (TRENCAM, hereafter TC), a synthetic mimic of Ent, to facilitate drug delivery across the outer membrane (OM) of Gram-negative pathogens. Conjugation of Amp to a new monofunctionalized TC scaffold affords TC-Amp, which displays markedly enhanced antibacterial activity against the gastrointestinal pathogen Salmonella enterica serovar Typhimurium ( S Tm) compared with unmodified Amp. Bacterial uptake, antibiotic susceptibility, and microscopy studies with S Tm show that the TC moiety facilitates TC-Amp uptake by the OM receptors FepA and IroN and that the Amp warhead inhibits penicillin-binding proteins. Moreover, TC-Amp achieves targeted activity, selectively killing S Tm in the presence of a commensal lactobacillus. Remarkably, we uncover that TC-Amp and its Ent-based predecessor Ent-Amp achieve enhanced antibacterial activity against diverse Gram-negative ESKAPE pathogens that express Ent uptake machinery, including strains that possess intrinsic β-lactam resistance. TC-Amp and Ent-Amp exhibit potency comparable to that of the FDA-approved SAC cefiderocol against Gram-negative pathogens. These results demonstrate the effective application of native and appropriately designed nonnative siderophores as vectors for drug delivery across the OM of multiple Gram-negative bacterial pathogens.

The antibiotic cefiderocol hijacks iron transporters to facilitate its uptake and resists β-lactamase degradation. While effective, resistance has been detected clinically with unknown mechanisms. Here, using experimental evolution, we identified cefiderocol resistance mutations in Pseudomonas aeruginosa. Resistance was multifactorial in host-mimicking growth media, led to multidrug resistance and paid fitness costs in cefiderocol-free environments. However, kin selection drove some resistant populations to cross-protect susceptible individuals from killing by increasing pyoverdine secretion via a two-component sensor mutation. While pyochelin sensitized P. aeruginosa to cefiderocol killing, pyoverdine and the enterobacteria siderophore enterobactin displaced iron from cefiderocol, preventing uptake by susceptible cells. Among 113 P. aeruginosa intensive care unit clinical isolates, pyoverdine production directly correlated with cefiderocol tolerance, and high pyoverdine producing isolates cross-protected susceptible P. aeruginosa and other Gram-negative bacteria. These in vitro data show that antibiotic cross-protection can occur via degradation-independent mechanisms and siderophores can serve unexpected protective cooperative roles in polymicrobial communities., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

In contaminated water and soil, little is known about the role and mechanism of the biometabolic molecule siderophore desferrioxamine-B (DFO) in the biogeochemical cycle of uranium due to complicated coordination and reaction networks. Here, a joint experimental and quantum chemical investigation is carried out to probe the biomineralization of uranyl (UO 2 2+ , referred to as U(VI) hereafter) induced by Shewanella putrefaciens (abbreviated as S. putrefaciens ) in the presence of DFO and Fe 3+ ion. The results show that the production of mineralized solids {hydrogen-uranium mica [H 2 (UO 2 ) 2 (PO 4 ) 2 ·8H 2 O]} via S. putrefaciens binding with UO 2 2+ is inhibited by DFO, which can both chelate preferentially UO 2 2+ to form a U(VI)-DFO complex in solution and seize it from U(VI)-biominerals upon solvation. However, with Fe 3+ ion introduced, the strong specificity of DFO binding with Fe 3+ causes re-emergence of biomineralization of UO 2 2+ {bassetite [Fe(UO 2 ) 2 (PO 4 ) 2 ·8(H 2 O)]} by S. putrefaciens , owing to competitive complexation between Fe 3+ and UO 2 2+ for DFO. As DFO possesses three hydroxamic functional groups, it forms hexadentate coordination with Fe 3+ and UO 2 2+ ions via these functional groups. The stability of the Fe 3+ -DFO complex is much higher than that of U(VI)-DFO, resulting in some DFO-released UO 2 2+ to be remobilized by S. putrefaciens . Our finding not only adds to the understanding of the fate of toxic U(VI)-containing substances in the environment and biogeochemical cycles in the future but also suggests the promising potential of utilizing functionalized DFO ligands for uranium processing.

Blocking iron uptake and metabolism has been emerging as a promising therapeutic strategy for the development of novel antimicrobial compounds. Like all mycobacteria, M. abscessus (Mab) has evolved several countermeasures to scavenge iron from host carrier proteins, including the production of siderophores, which play a crucial role in these processes. In this study, we solved, for the first time, the crystal structure of Mab-SaS, the first enzyme involved in the biosynthesis of siderophores. Moreover, we screened a small, focused library and identified a compound exhibiting a potent inhibitory effect against Mab-SaS (IC 50  ≈ 2 μM). Its binding mode was investigated by means of Induced Fit Docking simulations, performed on the crystal structure presented herein. Furthermore, cytotoxicity data and pharmacokinetic predictions revealed the safety and drug-likeness of this class of compounds. Finally, the crystallographic data were used to optimize the model for future virtual screening campaigns. Taken together, the findings of our study pave the way for the identification of potent Mab-SaS inhibitors, based on both established and unexplored chemotypes., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Laurent R. Chiarelli reports financial support was provided by Italian Cystic Fibrosis Research Foundation (FFC Ricerca). If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Aims: The study aims to explore antifungal properties of bacillibactin siderophore produced by the plant growth-promoting rhizobacterium (PGPR) Bacillus subtilis against fungal phytopathogens Alternaria porri and Fusarium equiseti isolated from Solanum lycopersicum and Solanum melongena plants., Methods and Results: Alternaria porri and F. equiseti were isolated from infected plants of eggplant and tomato, respectively. A plate assay was employed to assess the effect of bacillibactin against the phytopathogens. The antifungal potential of the PGPR was evaluated by estimation of dry fungal biomass, visualization of cellular deformity using compound and scanning electron microscopy, antioxidative enzyme assay and analysis of membrane damage via using lipid peroxidation. Inductively coupled plasma atomic emission spectroscopy (ICP-AES) analysis was employed to investigate changes in intracellular iron content. The impact of bacillibactin on pathogenesis was evaluated by infecting detached leaves of S. lycopersicum and S. melongena plants with both the pathogens and treating the infected leaves with bacillibactin. Leaves were further investigated for ROS accumulation, extent of necrosis and cell death. Our findings revealed significant damage to the hyphal structure of A. porri and F. equiseti following treatment with bacillibactin. Biomass reduction, elevated antioxidative enzyme levels, and membrane damage further substantiated the inhibitory effects of the siderophore on fungal growth. ICP-AES analysis indicates an increase in intracellular iron content suggesting enhanced iron uptake facilitated by bacillibactin. Moreover, application of 1500 µg ml-1 bacillibactin on infected leaves demonstrated a substantial inhibition of ROS accumulation, necrosis, and cell death upon bacillibactin treatment., Conclusions: This study confirms the potent antagonistic activity of bacillibactin against both the phytopathogens A. porri and F. equiseti growth, supporting its potential as a promising biological control agent for fungal plant diseases. Bacillibactin-induced morphological, physiological, and biochemical alterations in the isolated fungi and pathogen-infected leaves highlight the prospects of bacillibactin as an effective and sustainable solution to mitigate economic losses associated with fungal infections in vegetable crops., (© The Author(s) 2024. Published by Oxford University Press on behalf of Applied Microbiology International.)

The human skin microbiome consists of many species of bacteria, including Staphylococcus aureus and S. epidermidis. Individuals with atopic dermatitis (AD) have an increased relative abundance of S. aureus, which exacerbates the inflammation of AD. Although S. epidermidis, a main component of healthy skin microbiota, inhibits the growth of S. aureus, the balance between S. epidermidis and S. aureus is disrupted in the skin of individuals with AD. In this study, we found that Citrobacter koseri isolated from patients with AD produces substances that inhibit the growth of S. epidermidis. Heat-treated culture supernatant (CS) of C. koseri inhibited the growth of S. epidermidis but not S. aureus. The genome of C. koseri has gene clusters related to siderophores and the heat-treated CS of C. koseri contained a high concentration of siderophores compared with the control medium. The inhibitory activity of C. koseri CS against the growth of S. epidermidis was decreased by the addition of iron, but not copper or zinc. Deferoxamine, an iron-chelating agent, also inhibited the growth of S. epidermidis, but not that of S. aureus. These findings suggest that C. koseri inhibits the growth of S. epidermidis by interfering with its iron utilization., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Antimicrobial resistance is currently recognized as an urgent concern against public health in worldwide. Carbapenem-resistant (CR) Gram-negative bacteria, such as Enterobacterales, Pseudomonas aeruginosa and Acinetobacter baumannii are listed as critical pathogens which are widely spread and can cause severe and often deadly infections in WHO guidance. Cefiderocol (Fetroja ® ), a novel and first siderophore cephalosporin, was approved for the infections caused by these problematic CR Gram-negative bacteria in Japan on November 30, 2023. Cefiderocol has unique mechanisms to be incorporated into bacterial cells using bacterial iron transportation system and to be highly stable against most β-lactamases, which lead to promising antibacterial activity against these Gram-negative bacteria including CR strains in vitro. In CREDIBLE-CR Ph3 trial, cefiderocol showed the good efficacy and safety for patients with CR Gram-negative bacteria. In APEKS-cUTI and APEKS-NP trials, cefiderocol showed non-inferiority and suggested superiority to imipenem/cilastatin in complicated urinary tract infection (cUTI) patients, and non-inferiority to high dose of meropemen in pneumonia patients, respectively. Cefiderocol is expected to be an optimal treatment for CR Gram-negative infections with limited treatment options and would be an important drug to combat the threat of CR bacteria.

Siderophore-antibiotic conjugates (SACs) are of past and current interest for delivering antibacterials into Gram-negative bacterial pathogens that express siderophore receptors. Studies of SACs are often multifaceted and involve chemical and biological approaches. Major goals are to evaluate the antimicrobial activity and uptake of novel SACs and use the resulting data to inform further mode-of-action studies and molecular design strategies. In this chapter, we describe four key methods that we apply when investigating the antimicrobial activity and uptake of novel SACs based on the siderophore enterobactin (Ent). These methods are based on approaches from the siderophore literature as well as established protocols for antimicrobial activity testing, and include assays for evaluating SAC antimicrobial activity, time-kill kinetics, siderophore competition, and bacterial cell uptake using 57 Fe. These assays have served us well in characterizing our Ent-based conjugates and can be applied to study SACs that use other siderophores as targeting vectors., (Copyright © 2024. Published by Elsevier Inc.)

Siderophores are low-molecular-mass, high-affinity chelators of Fe 3+ ions that are critical for the survival of bacteria in ferric deficient environment. Exogenous siderophores are potential bacteriostat by disrupting the iron-uptake process of pathogens. In our previous work to discover siderophores, strain LS1784 was previously predicted to produce new catecholate-type siderophores by genome analysis but no compounds were obtained. In this work, we reclassified train LS1784 as Kitasatospora sp. LS1784 according to the genome phylogenetic analysis. Then guided by CAS colorimetric assay and molecular network analysis, four catecholate-type siderophores were isolated from the ethyl acetate extract of LS1784 which were coincident with the initial prediction. Notably, compounds 2 and 3 were reported for the first time. Following activity screening, compound 3 showed sufficient anti-Pseudomonas aeruginosa-infection activity in Caenorhabditis elegans infection models, whereas all compounds exhibited no antimicrobial activity. These results indicated that compound 3 can enhance the survival of P. aeruginosa infecting C. elegans by reducing the virulence of P. aeruginosa rather than killing P. aeruginosa, which aligns with our previous findings. Moreover, these findings highlight the effectiveness of comprehensive approaches, including genome mining, CAS (Chromeazurol S) testing, and molecular network (MN) analysis, in identifying potential siderophores, thereby expanding the siderophores arsenal in bacteria for the development of anti-infective drugs., (© 2023. The Author(s), under exclusive licence to the Japan Antibiotics Research Association.)

Antibacterial drug resistance of gram-negative bacteria (GNB) results in high morbidity and mortality of GNB infection, seriously threaten human health globally. Developing new antibiotics has become the critical need for dealing with drug-resistant bacterial infections. Cefiderocol is an iron carrier cephalosporin that achieves drug accumulation through a unique "Trojan horse" strategy into the bacterial periplasm. It shows high antibacterial activity against multidrug-resistant (MDR) Enterobacteriaceae and MDR non-fermentative bacteria. The application of cefiderocol offers new hope for treating clinical drug-resistant bacterial infections. However, limited clinical data and uncertainties about its resistance mechanisms constrain the choice of its therapeutic use. This review aimed to summarize the clinical applications, drug resistance mechanisms, and co-administration of cefiderocol., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Background: Cefiderocol (CFDC) is a novel siderophore-cephalosporin, which usually penetrates the bacteria through the iron-uptake pathways. Data is limited on the factors affecting CFDC activity and methods for overcoming resistance development. Synergistic approaches are needed to tackle antimicrobial resistance. This study aimed to determine CFDC activity on Klebsiella pneumoniae isolates from patients attending a single hospital in the United Arab Emirates (UAE), to explore the effect of β-lactamases on CFDC activity and to enhance CFDC susceptibility in both iron-depleted and iron-enriched conditions., Methods: We investigated 238 K. pneumoniae strains from diverse clinical sources. β-lactamase genes were detected by PCR. Susceptibility to CFDC and 12 comparator antibiotics were tested. Combinations of CFDC with β-lactamase inhibitors (BLIs) and/or an outer membrane (OM) permeabilizer (polymyxin B nonapeptide) were tested in iron-depleted and iron-enriched conditions., Results: CFDC exhibited efficacy of 97.9%, against multidrug-resistant (MDR), and extensively drug-resistant (XDR) strains, in addition to strains resistant to the last resort drugs such as colistin and tigecycline, including dual carbapenemase-producers (bla NDM and bla OXA-48-like ) with MIC ≤ 0.06-8 µg/ml. It was effective in killing strains with single and multiple β-lactamases; however, it lost activity in iron-enriched conditions. Synergy was achieved with dual combination of CFDC and BLIs, especially avibactam, which caused a significant reduction in MICs even in iron-enriched conditions. A significant reduction was seen with the triple combination including an OM permeabilizer plus avibactam. Killing-kinetic studies proved that the combination therapy caused dose reduction and faster killing by CFDC than the monotherapy., Conclusions: CFDC was deemed effective against MDR and XDR K. pneumoniae. Synergistic combination of CFDC with BLIs and OM permeabilizers could be effective to treat infections in iron-rich sites, but this should be investigated in vivo., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Farah Al-Marzooq reports financial support was provided by United Arab Emirates University. Farah Al-Marzooq reports a relationship with United Arab Emirates University that includes: funding grants., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

SbnE is an essential enzyme for staphyloferrin B biosynthesis in Staphylococcus aureus. An earlier study showed that natural product baulamycin A has in vitro inhibitory activity against SbnE and antibacterial potency. A SAR study with analogues of baulamycin A was conducted to identify potent inhibitors of SbnE and/or effective antibiotics against MRSA. The results show that selected analogues, including 11, 18, 21, 24a, 24c, 24m and 24n, exhibit single-digit micromolar inhibitory potencies against SbnE (IC 50 s = 1.81-8.94 μM) and 11, 24m, 24n possess significant activities against both SbnE (IC 50 s = 4.12-6.12 μM) and bacteria (MICs = 4-32 μg/mL). Biological investigations revealed that these substances possess potent cell wall disruptive activities and that they inhibit siderophore production in MRSA. Among the selected analogues, 7 has excellent antibiotic activities both gram-positive and -negative bacteria (0.5-4 μg/mL). Moreover, these analogues significantly impede biofilm formation in a concentration-dependent manner. Taken together, the results of the investigation provide valuable insight into the nature of novel baulamycin A analogues that have potential efficacy against MRSA owing to their membrane damaging activity and/or inhibitory efficacy against siderophore production., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Taebo Sim is a shareholder of Magicbullettherapeutics Inc., (Copyright © 2023 Elsevier Masson SAS. All rights reserved.)

Cefiderocol (CFDC) is the first-in-class siderophore-cephalosporin. Klebsiella pneumoniae strain that is extremely resistant to CFDC (MIC: 256 µg/ml) was isolated for the first time in the United Arab Emirates from a patient with pneumonia and sepsis. It belonged to sequence-type 14 (ST14), with a novel core genome ST. Resistance was driven by the co-expression of β-lactamases (bla NDM-1 , bla OXA-232 and bla CTX-M-15 ) and a mutation in catecholate-siderophore receptor, utilized by CFDC to enter the bacterial cell. Synergistic combinations (β-lactamase inhibitors, aztreonam plus CFDC) re-sensitized the bacteria to CFDC. Although CFDC resistance is multifactorial, the combination with β-lactamase inhibitors represents a promising approach in resistance reversal for fighting superbugs., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The natural prenylated chalcone isobavachalcone (IBC) shows good antibacterial activity against Gram-positive bacteria but is ineffective against Gram-negative bacteria, most likely due to the outer membrane barrier of Gram-negative bacteria. The Trojan horse strategy has been shown to be an effective strategy to overcome the reduction in the permeability of the outer membrane of Gram-negative bacteria. In this study, eight different 3-hydroxy-pyridin-4(1H)-one-isobavachalcone conjugates were designed and synthesized based on the siderophore Trojan horse strategy. The conjugates exhibited 8- to 32-fold lower minimum inhibitory concentrations (MICs) and 32- to 177-fold lower half-inhibitory concentrations (IC 50 s) against Pseudomonas aeruginosa PAO1 as well as clinical multidrug-resistant (MDR) strains compared to the parent IBC under iron limitation. Further studies showed that the antibacterial activity of the conjugates was regulated by the bacterial iron uptake pathway under different iron concentration conditions. Studies on the antibacterial mechanism of conjugate 1b showed that it exerts antibacterial activity by disrupting cytoplasmic membrane integrity and inhibiting cell metabolism. Finally, conjugate 1b showed a lower cytotoxic effects on Vero cells than IBC and a positive therapeutic effect in the treatment of bacterial infections caused by Gram-negative bacteria PAO1. Overall, this work demonstrates that IBC can be delivered to Gram-negative bacteria when combined with 3-hydroxy-pyridin-4(1H)-ones as siderophores and provides a scientific basis for the development of effective antibacterial agents against Gram-negative bacteria., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Masson SAS. All rights reserved.)

The search and creation of innovative antimicrobial drugs, acting against resistant and multiresistant strains of bacteria and fungi, are one of the most important tasks of modern bioorganic chemistry and pharmaceuticals. Since iron is essential for the vital activity of almost all organisms, including mammals and bacteria, the proteins involved in its metabolism can serve as potential targets in the development of new promising antimicrobial agents. Such targets include endogenous mammalian biomolecules, heme oxygenases, siderophores, protein 24p3, as well as bacterial heme oxygenases and siderophores. Other proteins that are responsible for the delivery of iron to cells and its balance between bacteria and the host organism also attract certain particular interest. The review summarizes data on the development of inhibitors and inducers (activators) of heme oxygenases, selective for mammals and bacteria, and considers the characteristic features of their mechanisms of action and structure. Based on the reviewed literature data, it was concluded that the use of hemin, the most powerful hemooxygenase inducer, and its derivatives as potential antimicrobial and antiviral agents, in particular against COVID-19 and other dangerous infections, would be a promising approach. In this case, an important role is attributed to the products of hemin degradation formed by heme oxygenases in vitro and in vivo. Certain attention has been paid to the data on the antimicrobial action of iron-free protoporphyrinates, namely complexes with Co, Ga, Zn, Mn, their advantages and disadvantages compared to hemin. Modification of the well-known antibiotic ceftazidime with a siderophore molecule increased its effectiveness against resistant bacteria.

Intravenous gallium therapy is a non-antibiotic approach to limit Pseudomonas aeruginosa biofilm proliferation, by outcompeting iron for siderophore binding. Gallium therapy represents a viable therapeutic strategy for cystic fibrosis (CF) patients harbouring mucoid P. aeruginosa biofilm lung infections. Siderophore deficient P. aeruginosa isolates still demonstrate a hindered biofilm proliferation when exposed to gallium but it is currently unknown whether exogenous gallium has any disruptive influence on the exopolysaccharide (EPS), the major mucoid P. aeruginosa CF lung biofilm matrix component. To that end, Density-Functional Theory (DFT) was deployed to assess whether gallium (Ga3+) could be substituted into the mature mucoid EPS scaffold in preference of calcium (Ca2+)-the native EPS cross-linking ion. Removal of the stable, bound native calcium ions offers a large enthalpic barrier to the substitution and the mature EPS fails to accommodate exogenous gallium. This suggests that gallium, perhaps, is utilising a novel, possibly unknown, ferric uptake system to gain entry to siderophore deficient cells., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Hills et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Deferoxamine (DFB) is a trihydroxamic acid siderophore that chelates with iron (Fe) to form iron-siderophore complexes. The existence of siderophores in nature changes the form of iron and affects the absorption and utilization of iron by organisms. However, the relationship between siderophores and the growth of Cyanobacteria is largely unknown. In this study, the cellular and transcriptomic responses to the addition of DFB were investigated. A high concentration of DFB (12 mg/L) significantly inhibited the growth of Cyanobacteria cells, reduced photosynthetic activity, and induced the production of peroxidase, with the highest inhibition rate of algal growth of 74.82%. These indexes were also affected for the low (3 mg/L) and medium concentration (6 mg/L) groups, but this difference is closely related to the growth stage of Cyanobacteria cells. This may be due to competition between the cell-associated iron-binding part/system and the extracellular Fe (Ⅲ)-DFB ligand. Transcriptome results showed that most of the genes involved in iron uptake and transport were down-regulated, and only the fur gene encoding the iron uptake regulator protein was significantly up-regulated. Most genes related to photosynthesis, glycolysis, and fatty acid metabolism were also down-regulated, while the obvious up-regulation of a few genes may be a complex regulation in response to the down-regulation of most genes. These findings will provide important insights into the effects of siderophores on iron bioavailability in algae., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Emergence of cefiderocol resistance among carbapenemase-producing Enterobacterales, particularly those in the Enterobacter cloacae complex (ECC), is becoming of alarming concern; however, the mechanistic basis of this phenomenon remains poorly understood. We describe the acquisition of VIM-1-mediated reduced cefiderocol susceptibility (MICs 0.5 to 4 mg/L) in a collection of 54 carbapenemase-producing isolates belonging to the ECC. MICs were determined by reference methodologies. Antimicrobial resistance genomic analysis was performed through hybrid WGS. The impact of VIM-1 production on cefiderocol resistance in the ECC background was examined at microbiological, molecular, biochemical, and atomic levels. Antimicrobial susceptibility testing yielded 83.3% susceptible isolates and MIC 50/90 values of 1/4 mg/L. Decreased susceptibility to cefiderocol was mainly associated with isolates producing VIM-1, with cefiderocol MICs 2- to 4-fold higher than for isolates carrying other types of carbapenemases. E. cloacae and Escherichia coli VIM-1 transformants displayed significantly enhanced cefiderocol MICs. Biochemical assays with purified VIM-1 protein revealed low but detectable cefiderocol hydrolysis. Simulation studies revealed how cefiderocol is anchored to the VIM-1 active site. Additional molecular assays and WGS data analysis highlighted the implication of SHV-12 coproduction and suggested the inactivation of the FcuA-like siderophore receptor as further contributors to the higher cefiderocol MICs. Our findings warn of the potential of the VIM-1 carbapenemase to at least partly limit the activity of cefiderocol in the ECC. This effect is probably enhanced due to combination with additional mechanisms, such as ESBL production and siderophore inactivation, and indicates the need for active surveillance to extend the life span of this promising cephalosporin., Competing Interests: The authors declare no conflict of interest.

Knowledge on resistance mechanisms toward cefiderocol, a novel siderophore-conjugated cephalosporin antibiotic, is still limited. Although the presence of New-Delhi metallo-β-lactamase has been demonstrated to facilitate the resistance development toward cefiderocol via siderophore receptor mutations in Enterobacter cloacae and Klebsiella pneumoniae, the impact of metallo-β-lactamases on facilitating such mutations in Escherichia coli is not yet elucidated. Our study aimed to study the effect of the presence of various β-lactamases, such as NDM-5, VIM-1, KPC-2, and OXA-48, on the development of cefiderocol resistance in E. coli. To this end, we performed liquid mating to transfer these β-lactamases onto a defined K-12 E. coli background (J53) and exposed these transconjugants to increasing cefiderocol concentrations in a serial passage experiment. Cefiderocol-resistant isolates were genotyped by whole-genome sequencing to investigate the underlying resistance mechanism. Cefiderocol-resistant isolates emerged only in isolates producing VIM-1 and NDM-5 metallo-β-lactamase, but not in those producing the serine β-lactamases KPC-2 and OXA-48. We observed two distinct morphological changes of the J53 E. coli strain exhibiting reduced colony size after insertions of transposable elements in the tonB gene leading to alterations in the TonB binding site and morphological changes consistent with the small-colony variant (SCV) phenotype due to mutations in the hemB and hemH genes. Passaging experiments suggested that these phenotypes were highly plastic. The SCV phenotype is attributed to immune evasion and decreased susceptibility toward antibiotics. The emergence of SCV following cefiderocol exposure may have clinical implications for bacterial clearance and warrants further investigation., Competing Interests: The authors declare a conflict of interest. Dennis Nurjadi received speaker honorarium for Webcast from Shionogi.

Burkholderia pyrrocinia JK-SH007 can effectively control poplar canker caused by pathogenic fungi. Its antifungal mechanism remains to be explored. Here, we characterized the functional role of CysB in B. pyrrocinia JK-SH007. This protein was shown to be responsible for the synthesis of cysteine and the siderophore ornibactin, as well as the antifungal activity of B. pyrrocinia JK-SH007. We found that deletion of the cysB gene reduced the antifungal activity and production of the siderophore ornibactin in B. pyrrocinia JK-SH007. However, supplementation with cysteine largely restored these two abilities in the mutant. Further global transcriptome analysis demonstrated that the amino acid metabolic pathway was significantly affected and that some sRNAs were significantly upregulated and targeted the iron-sulfur metabolic pathway by TargetRNA2 prediction. Therefore, we suggest that, in B. pyrrocinia JK-SH007, CysB can regulate the expression of genes related to Fe-S clusters in the iron-sulfur metabolic pathway to affect the antifungal activity of B. pyrrocinia JK-SH007. These findings provide new insights into the various biological functions regulated by CysB in B. pyrrocinia JK-SH007 and the relationship between iron-sulfur metabolic pathways and fungal inhibitory substances. Additionally, they lay the foundation for further investigation of the main antagonistic substances of B. pyrrocinia JK-SH007.

During chronic cystic fibrosis (CF) infections, evolved Pseudomonas aeruginosa antibiotic resistance is linked to increased pulmonary exacerbations, decreased lung function, and hospitalizations. However, the virulence mechanisms underlying worse outcomes caused by antibiotic resistant infections are poorly understood. Here, we investigated evolved aztreonam resistant P. aeruginosa virulence mechanisms. Using a macrophage infection model combined with genomic and transcriptomic analyses, we show that a compensatory mutation in the rne gene, encoding RNase E, increased pyoverdine and pyochelin siderophore gene expression, causing macrophage ferroptosis and lysis. We show that iron-bound pyochelin was sufficient to cause macrophage ferroptosis and lysis, however, apo-pyochelin, iron-bound pyoverdine, or apo-pyoverdine were insufficient to kill macrophages. Macrophage killing could be eliminated by treatment with the iron mimetic gallium. RNase E variants were abundant in clinical isolates, and CF sputum gene expression data show that clinical isolates phenocopied RNase E variant functions during macrophage infection. Together these data show how P. aeruginosa RNase E variants can cause host damage via increased siderophore production and host cell ferroptosis but may also be targets for gallium precision therapy., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Vaillancourt et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Aim: The present study aimed to investigate the effect of bacterivorous soil protists on plant growth promoting (PGP) attributes of bacterial species and their co-inoculative impact on rice seedling growth., Methods and Results: The effect of protists on the PGP attributes of bacteria was tested using standard protocols. The results revealed that the plant-beneficial properties of plant growth promoting bacteria (PGPB) were altered in the presence of various protist species. A significant increase in the production of siderophore units (86.66%), ammonia (34.80 μmol mL-1), and phosphate solubilization index (PSI) (5.6) was observed when Bacillus cereus (Bc) and Pseudomonas fluorescens (Ps) were co-inoculated with unidentified species belonging to the family Kreyellidae (C5). In the case of Enterobacter cloacae co-inoculated with C5 (Kreyellidae), a higher amount of siderophore (51.33%), ammonia (25.18 μmol mL-1), and indole-3-acetic acid (IAA)-like substance (28.59 μg mL-1) production were observed. The biofilm-forming ability of B. cereus is enhanced in the presence of Tetrahymena sp. (C2Bc), unidentified Kreyellidae (C5Bc), and Colpoda elliotti (C12Bc), whereas E. cloacae showed higher biofilm formation in the presence of Tetrahymena sp. alone Although IAA production decreased under predation pressure, a significant increase in shoot length (64.24%) and primary root length (98.18%) in co-inoculative treatments (C12Bc and C5Bc) compared to bacteria alone (25% and 61.50% for shoots and roots, respectively) was observed. The results of enhanced PGP attributes and rice seedlings growth under predation pressure correlated with the enhanced bacterial activity under predation pressure and protist involvement in plant growth development., Conclusions: Protists may act as regulators of the bacterial activities involved in plant growth promotion and thus enhance plant growth., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

Background: Intracerebral hemorrhage (ICH) is a stroke with a high morbidity and mortality rate. Deferoxamine (DFX) is thought to be effective in treating Intracerebral Hemorrhage. In our study, we performed a meta-analysis to evaluate the treatment effects of DFX., Methods: We systematically searched PubMed, Embase, Web of Science, the Cochrane Central Register of Controlled Trials, and Chinese Biomedical Literature Database in Jan 2022 for studies on DFX for ICH patients. Outcome measures included relative hematoma volume, relative edema volume, good neurological functional outcome and adverse events. Odds risk (OR) and weighted mean difference (WMD) were used to evaluate clinical outcomes., Results: After searching 636 articles, 4 RCTs, 2 NRCTs, and 1cohort study were included. We found that DFX was effective in hematoma absorption on day 7 after onset, but the difference was not significant on day 14. DFX could suppress edema expansion on days 3, 7, and 14 after onset. DFX did not contribute to better outcomes after 3 and 6 months when used the modified Rankin Scale and the Glasgow Outcome Scale to evaluate neurological prognosis. The pooled results showed no statistically significant difference in Serious adverse events between the experimental and control groups., Conclusions: DFX could limit edema expansion on days 3, 7, and 14 after commencement and facilitate hematoma absorption at week 1 without significantly increasing the risk of adverse events, but it did not improve neurological prognosis., Competing Interests: Declaration of Competing Interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Despite advances in chemotherapeutic interventions for the treatment of malaria, there is a continuing need for the development of new antimalarial agents. Previous studies indicated that co-administration of chloroquine with antioxidants such as the iron chelator deferoxamine (DFO) prevented the development of persistent cognitive damage in surrogate models of cerebral malaria. The work described herein reports the syntheses and antimalarial activities of covalent conjugates of both natural (siderophores) and artificial iron chelators, namely DFO, ferricrocin and ICL-670, with antimalarial 1,2,4-trioxolanes (ozonides). All of the synthesized conjugates had potent antimalarial activities against the in vitro cultures of drug resistant and drug sensitive strains of Plasmodium falciparum. The work described herein provides the basis for future development of covalent combination of iron chelators and antimalarial chemotherapeutic agents for the treatment of cerebral malaria., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Achieving cellular uptake is a central challenge for novel antibiotics targeting Gram-negative bacterial pathogens. One strategy is to hijack the bacterial iron transport system by siderophore-antibiotic conjugates that are actively imported into the cell. This was realized with the MECAM-ampicillin conjugate LP-600 we recently reported that was highly active against E. coli . In the present study, we investigate a paradoxical regrowth of E. coli upon treatment of LP-600 at concentrations 16-32 times above the minimum inhibitory concentration (MIC). The phenomenon, coined "Eagle-effect" in other systems, was not due to resistance formation, and it occurred for the siderophore conjugate but not for free ampicillin. To investigate the molecular imprint of the Eagle effect, a combined transcriptome and untargeted metabolome analysis was conducted. LP-600 induced the expression of genes involved in iron acquisition, SOS response, and the e14 prophage upon regrowth conditions. The Eagle effect was diminished in the presence of sulbactam, which we ascribe to a putative synergistic antibiotic action but not to β-lactamase inhibition. The study highlights the relevance of the Eagle effect for siderophore conjugates. Through the first systematic -omics investigations, it also demonstrates that the Eagle effect manifests not only in a paradoxical growth but also in unique gene expression and metabolite profiles.

The bacterial endophytes isolated from the halophyte Salicornia brachiata were explored for the antimicrobial potential to discover novel microbial inhibitors that combat multidrug resistance. Upon investigation, ethyl acetate extract of the endophyte Bacillus subtilis NPROOT3 displayed significant potency against Mycobacterium smegmatis MTCC6 as well as Mycobacterium tuberculosis H37Rv strain. Further investigation of ethyl acetate crude extract by repeated chromatographic separations followed by characterization using UV, HR-ESI-MS, MALDI-MS, MALDI-MS/MS, CD, and NMR spectroscopy yielded a series of five known siderophores, namely, SVK21 (1), bacillibactin C (2), bacillibactin B (3), tribenglthin A (4), and bacillibactin (5). A total of two out of five compounds, 4 (MIC 38.66 μM) and 5 (MIC 22.15 μM) exhibited significant inhibition against the strain M. smegmatis MTCC6 comparable with positive control rifampicin (MIC 12.15 μM). None of these five bacillibactin molecules are previously reported to exhibit bioactivity against Mycobacterium sp. Herein for the first time, all the compounds were screened for their antibacterial activities against a panel of bacterial pathogens of humans. Furthermore, the probable mechanism of action of bacillibactin compounds for their antimycobacterial activity is also discussed. The findings of this study open up a new chemotype for inhibition of the Mycobacterium sp. and other multidrug-resistant pathogens., (© The Author(s) 2022. Published by Oxford University Press on behalf of Applied Microbiology International.)

Antibacterial resistance is a healthcare burden. Among Gram-negative bacteria, Pseudomonas aeruginosa belongs to the first list of antibiotic-resistant "priority pathogens" described by the World Health Organization. Formerly Pseudomonas pseudomallei, Burkholderia pseudomallei, responsible for melioidosis, is considered as a potential bioterrorist weapon by the Centers of Diseases Control and Prevention. We are interested in the development of new ways to combat these bacteria, targeted due to their high level of resistance to antibiotics via a lack of membrane permeability or efflux. Using iron transport systems is a promising strategy to bypass the bacteria cell membrane and restore the activity of conventional antibiotics such as ciprofloxacin. Specific outer membrane receptors are necessary to most microbes as they allow iron uptake, essential for their survival through siderophore-dependent mechanisms. These systems may allow the introduction of antibacterial agents, chemically coupled to a natural or synthetic siderophore molecule to form siderophore-antibiotic conjugates. In this work, we describe the synthesis of six new siderophore analog-ciprofloxacin conjugates including cleavable linker or not. The siderophore analogs correspond to a mono-catechol or a hydroxypyridinone moiety recognized by both Pseudomonas and Burkholderia species. Physico-chemical studies showed that (i) conjugates were unable to interact or cross the membrane by passive diffusion and (ii) conjugates with cleavable linker are stable in physiologic environment. Biological evaluations have highlighted a promising compound 2d, bearing an hydroxypyridinone moiety with a cleavable linker, active on a large panel of strains of Pseudomonas aeruginosa, Burkholderia pseudomallei and Burkholderia thailandensis without toxicity observed in vitro., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Masson SAS. All rights reserved.)

Background: Cefiderocol (CFDC) is a novel siderophore-cephalosporin, effective against multidrug-resistant Gram-negative bacteria. As it has a siderophore side chain, it can utilize iron acquisition systems for penetration of the bacterial outer membrane. We aimed to elucidate the role of siderophores and iron uptake receptors in defining Klebsiella pneumoniae susceptibility to CFDC., Methods: Initially, 103 K. pneumoniae strains were characterized for susceptibility to different antibiotics including CFDC. CFDC minimum inhibitory concentrations (MIC) were determined in iron-depleted and iron-enriched conditions. Iron uptake genes including siderophores, their receptors, ferric citrate (fecA) and iron uptake (kfu) receptors were detected by PCR in all the strains. For 10 selected strains, gene expression was tested in iron-depleted media with or without CFDC treatment and compared to expression in iron-enriched conditions., Results: CFDC exhibited 96.1% susceptibility, being superior to all the other antibiotics (MIC50: 0.5 and MIC90: 4 μg/ml). Only three strains (2.9%) were intermediately susceptible and a pandrug resistant strain (0.97%) was resistant to CFDC (MIC: 8 and 256 μg/ml, respectively). The presence of kfu and fecA had a significant impact on CFDC MIC, especially when co-produced, and if coupled with yersiniabactin receptor (fyuA). CFDC MICs were negatively correlated with enterobactin receptor (fepA) expression and positively correlated with expression of kfu and fecA. Thus, fepA was associated with increased susceptibility to CFDC, while kfu and fecA were associated with reduced susceptibility to CFDC. CFDC MICs increased significantly in iron-enriched media, with reduced expression of siderophore receptors, hence, causing less drug uptake., Conclusion: Iron acquisition systems have a significant impact on CFDC activity, and their altered expression is a factor leading to reduced susceptibility. Iron concentration is also a major player affecting CFDC susceptibility; therefore, it is essential to explore possible ways to improve the drug activity to facilitate its use to treat infections in iron-rich sites., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Daoud et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

This study aimed to investigate the antibacterial mechanism of cefiderocol (CFDC) using data-independent acquisition quantitative proteomics combined with cellular and molecular biological assays. Numerous differentially expressed proteins related to the production of NADH, reduced cofactor flavin adenine dinucleotide (FADH 2 ), NADPH and reactive oxygen species (ROS), iron-sulfur cluster binding, and iron ion homeostasis were found to be upregulated by CFDC. Furthermore, parallel reaction monitoring analysis validated these results. Meanwhile, we confirmed that the levels of NADH, ROS, H 2 O 2 , and iron ions were induced by CFDC, and the sensitivity of Escherichia coli to CFDC was inhibited by the antioxidant vitamin C, N -acetyl-l-cysteine, and deferoxamine. Moreover, deferoxamine also suppressed the H 2 O 2 stress induced by CFDC. In addition, knockout of the NADH-quinone oxidoreductase genes ( nuoA , nuoC , nuoE , nuoF , nuoG , nuoJ , nuoL , nuoM ) in the respiratory chain attenuated the sensitivity of E. coli to CFDC far beyond the effects of cefepime and ceftazidime; in particular, the E. coli BW25113 Δ nuoJ strain produced 60-fold increases in MIC to CFDC compared to that of the wild-type E. coli BW25113 strain. The present study revealed that CFDC exerts its antibacterial effects by inducing ROS stress by elevating the levels of NADH and iron ions in E. coli. IMPORTANCE CFDC was the first FDA-approved siderophore cephalosporin antibiotic in 2019 and is known for its Trojan horse tactics and broad antimicrobial activity against Gram-negative bacteria. However, its antibacterial mechanism is not fully understood, and whether it has an impact on in vivo iron ion homeostasis remains unknown. To comprehensively reveal the antibacterial mechanisms of CFDC, data-independent acquisition quantitative proteomics combined with cellular and molecular biological assays were performed in this study. The findings will further facilitate our understanding of the antibacterial mechanism of CFDC and may provide a theoretical foundation for controlling CFDC resistance in the future.

Biocontrol microbes are environment-friendly and safe for humans and animals. To seek biocontrol microbes effective in suppressing tomato gray mold is important for tomato production. Therefore, serial experiments were conducted to characterize the antagonism of Bacillus velezensis HY19, a novel self-isolated biocontrol bacterium, against Botrytis cinerea in vitro and the control on tomato gray mold in greenhouse. This bacterium produced extracellular phosphatase, protease, cellulose and siderophores, and considerably inhibited the growth of B. cinerea. A liquid chromatography-mass spectrometry (LC-MS) detected salicylic acid and numerous antifungal substances present in B. velezensis HY19 fermentation liquid (BVFL). When B. cinerea was grown on potato glucose agar, BVFL crude extract remarkably suppressed the fungal growth and reduced protein content and the activities of catalase (CAT), superoxide dismutase (SOD), and peroxidase (POD). Transcriptome studies showed that BVFL crude extract significantly induced different expression of numerous genes in B. cinerea, most of which were down-regulated. Theses differently expressed genes were involved in the biological process, cell compartment, molecular functions, and metabolisms of glycine, serine, threonine, and sulfur in pathogen hyphae. Thus, this biocontrol bacterium antagonized B. cinerea in multiple ways due to the production of numerous antifungal substances that acted on multiple targets in the cells. BVFL significantly increased antioxidant enzyme activities in tomato leaves and decreased the incidence of tomato gray mold, with the control efficacies of 73.12-76.51%. Taken together, B. velezensis HY19 showed a promising use potential as a powerful bioagent against tomato gray mold., Competing Interests: Declaration of Competing Interest The work described here was not published previously, and is not under consideration for publication elsewhere. There is no conflict of interest in the submission of this manuscript. The manuscript has been approved for publication by all authors., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Cefiderocol is a cephalosporin antibiotic presenting expanded antimicrobial activity. CirA is a gateway for cefiderocol to enter bacterial cells. We found that cirA1 and cirA198 were primary alleles in Klebsiella pneumoniae. CirA1 exhibited higher iron-transporting ability than CirA198 in iron-limited conditions. The cefiderocol minimum inhibitory concentration (MIC) increased from 0.5 mg/L to 2 mg/L when cirA1 was mutated to cirA198. Consistently, the MIC showed a 4-fold decrease when cirA198 was mutated to cirA1. Therefore, CirA1 has higher capacity to transport siderophores, contributing to increased cefiderocol susceptibility., Competing Interests: Declaration of Competing Interest None declared., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Potato production worldwide is plagued by several disease-causing pathogens that result in crop and economic losses estimated to billions of dollars each year. To this day, synthetic chemical applications remain the most widespread control strategy despite their negative effects on human and environmental health. Therefore, obtainment of superior biocontrol agents or their naturally produced metabolites to replace fungicides or to be integrated into practical pest management strategies has become one of the main targets in modern agriculture. Our main focus in the present study was to elucidate the antagonistic potential of a new strain identified as Bacillus subtilis EG21 against potato pathogens Phytophthora infestans and Rhizoctonia solani using several in vitro screening assays. Microscopic examination of the interaction between EG21 and R. solani showed extended damage in fungal mycelium, while EG21 metabolites displayed strong anti-oomycete and zoosporecidal effect on P. infestans . Mass spectrometry (MS) analysis revealed that EG21 produced antifungal and anti-oomycete cyclic lipopeptides surfactins (C 12 to C 19 ). Further characterization of EG21 confirmed its ability to produce siderophores and the extracellular lytic enzymes cellulase, pectinase and chitinase. The antifungal activity of EG21 cell-free culture filtrate (CF) was found to be stable at high-temperature/pressure treatment and extreme pH values and was not affected by proteinase K treatment. Disease-inhibiting effect of EG21 CF against P. infestans and R. solani infection was confirmed using potato leaves and tubers, respectively. Biotechnological applications of using microbial agents and their bioproducts for crop protection hold great promise to develop into effective, environment-friendly and sustainable biocontrol strategies. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Alternaria alternata botybirnavirus 1 (AaBRV1) was isolated from a strain of Alternaria alternata , causing watermelon leaf blight in our previous research. The effect of AaBRV1 on the phenotype of its host fungus, however, was not determined. In the present study, a novel strain of AaBRV1 was identified in A. tenuissima strain TJ-NH-51S-4, the causal agent of cotton Alternaria leaf spot, and designated as AaBRV1-AT1. A mycovirus AaBRV1-AT1-free strain TJ-NH-51S-4-VF was obtained by protoplast regeneration, which eliminated AaBRV1-AT1 from the mycovirus AaBRV1-AT1-infected strain TJ-NH-51S-4. Colony growth rate, spore production, and virulence of strain TJ-NH-51S-4 were greater than they were in TJ-NH-51S-4-VF, while the sensitivity of strain TJ-NH-51S-4 to difenoconazole, as measured by the EC 50 , was lower. AaBRV1-AT1 was capable of vertical transmission via asexual spores and horizontal transmission from strain TJ-NH-51S-4 to strain XJ-BZ-5-1 hyg (another strain of A. tenuissima ) through hyphal contact in pairing cultures. A total of 613 differentially expressed genes (DEGs) were identified in a comparative transcriptome analysis between TJ-NH-51S-4 and TJ-NH-51S-4-VF. Relative to strain TJ-NH-51S-4-VF, the number of up-regulated and down-regulated DEGs in strain TJ-NH-51S-4 was 286 and 327, respectively. Notably, the expression level of one DEG-encoding cytochrome P450 sterol 14α-demethylase and four DEGs encoding siderophore iron transporters were significantly up-regulated. To our knowledge, this is the first documentation of hypervirulence and reduced sensitivity to difenoconazole induced by AaBRV1-AT1 infection in A. tenuissima .