Siderophore-mediated Iron Uptake Research Articles

Growth of Pseudomonas aeruginosa in zinc poor environments is promoted by a nicotianamine-related metallophore.

Previous studies have suggested that P. aeruginosa possesses redundant zinc uptake systems. To identify uncharacterized zinc transporters, we analyzed the genome-wide transcriptional responses of P. aeruginosa PA14 to zinc restriction. This approach led to the identification of an operon (zrmABCD) regulated by the zinc uptake regulator Zur, that encodes for a metallophore-mediated zinc import system. This operon includes the genes for an uncharacterized TonB-dependent Outer Membrane Protein (ZrmA) and for a putative nicotianamine synthase (ZrmB). The simultaneous inactivation of the ZnuABC transporter and of one of these two genes markedly decreases the ability of P. aeruginosa to grow in zinc-poor media and compromises intracellular zinc accumulation. Our data demonstrate that ZrmB is involved in the synthesis of a metallophore which is released outside the cell and mediates zinc uptake through the ZrmA receptor. We also show that alterations in zinc homeostasis severely affect the ability of P. aeruginosa to cause acute lung and systemic infections in C57BL/6 mice, likely due to the involvement of zinc in the expression of several virulence traits. These findings disclose a hitherto unappreciated role of zinc in P. aeruginosa pathogenicity and reveal that this microorganism can obtain zinc through a strategy resembling siderophore-mediated iron uptake.

Investigation of a spontaneous mutant reveals novel features of iron uptake in Shewanella oneidensis

Shewanella oneidensis is among the first and the best studied bacteria capable of respiring minerals as terminal electron acceptors (EAs), including a variety of iron ores. This respiration process relies on a large number of c-type cytochromes, which per se are iron-containing proteins. Thus, iron plays an essential and special role in iron respiration of S. oneidensis, prompting extensive investigations into iron physiology. Despite this, we still know surprisingly little about the components and characteristics of iron transport in this bacterium. Here, we report that TonB-dependent receptor PutA (SO_3033) is specific to the siderophore-mediated iron uptake. Although homologs of PutA are abundant, none of them can function as a replacement. In the absence of PutA, S. oneidensis suffers from an iron shortage, which leads to a severe defect in production of cytochrome c. However, proteins requiring other types of cytochromes, such as b and d, do not appear to be significantly impacted. Intriguingly, lactate, but not other carbon sources that are routinely used to support growth, is able to promote iron uptake when PutA is missing. We further show that the lactate-mediated iron import is independent of lactate permeases. Overall, our results suggest that in S. oneidensis the siderophore-dependent pathway plays a key role in iron uptake when iron is limited, but many alternative routes exist.

TonB-Dependent Utilization of Dihydroxamate Xenosiderophores in Synechocystis sp. PCC 6803.

In Gram-negative bacteria, transport of ferric siderophores through outer membrane is a complex process that requires specific outer membrane transporters and energy-transducing TonB-ExbB-ExbD system in the cytoplasmic membrane. The genome of the non-siderophore-producing cyanobacterium Synechocystis sp. PCC 6803 encodes all putative components of the siderophore-mediated iron uptake system. So far, there has been no experimental evidence for the existence of such a pathway in this organism. On the contrary, its reductive iron uptake pathway has been studied in detail. We demonstrate that Synechocystis sp. PCC 6803 is capable of using dihydroxamate xenosiderophores, either ferric schizokinen (FeSK) or a siderophore of the filamentous cyanobacterium Anabaena variabilis ATCC 29413 (SAV), as the sole source of iron. Inactivation of the tonB gene or the exbB1-exbD1 gene cluster resulted in an inability to utilize these siderophores. At the same time, the inactivation of the feoB gene encoding FeoB plasma membrane ferrous iron transporter, or one of the futB or futC genes encoding permease and ATPase subunit of FutABC ferric iron transporter, did not impair the ability of cells to utilize FeSK or SAV as the sole source of iron for growth. Our data suggest that cyanobacterium Synechocystis sp. PCC 6803 is capable of acquiring iron-siderophore complexes in a TonB-dependent manner without iron reduction in the periplasm.

Aspergillusfumigatus protein phosphatase PpzA is involved in iron assimilation, secondary metabolite production, and virulence.

Metal restriction imposed by mammalian hosts during an infection is a common mechanism of defence to reduce or avoid the pathogen infection. Metals are essential for organism survival due to its involvement in several biological processes. Aspergillus fumigatus causes invasive aspergillosis, a disease that typically manifests in immunocompromised patients. A.fumigatus PpzA, the catalytic subunit of protein phosphatase Z (PPZ), has been recently identified as associated with iron assimilation. A.fumigatus has 2 high-affinity mechanisms of iron acquisition during infection: reductive iron assimilation and siderophore-mediated iron uptake. It has been shown that siderophore production is important for A.fumigatus virulence, differently to the reductive iron uptake system. Transcriptomic and proteomic comparisons between ∆ppzA and wild-type strains under iron starvation showed that PpzA has a broad influence on genes involved in secondary metabolism. Liquid chromatography-mass spectrometry under standard and iron starvation conditions confirmed that the ΔppzA mutant had reduced production of pyripyropene A, fumagillin, fumiquinazoline A, triacetyl-fusarinine C, and helvolic acid. The ΔppzA was shown to be avirulent in a neutropenic murine model of invasive pulmonary aspergillosis. PpzA plays an important role at the interface between iron starvation, regulation of SM production, and pathogenicity in A.fumigatus.

Differentially regulated high-affinity iron assimilation systems support growth of the various cell types in the dimorphic pathogen Talaromyces marneffei.

Iron is a key trace element important for many biochemical processes and its availability varies with the environment. For human pathogenic fungi iron acquisition can be particularly problematical because host cells sequester free iron as part of the acute-phase response to infection. Fungi rely on high-affinity iron uptake systems, such as reductive iron assimilation (RIA) and siderophore-mediated iron uptake (non-RIA). These have been extensively studied in pathogenic fungi that exist outside of host cells, but much less is known for intracellular fungal pathogens. Talaromyces marneffei is a dimorphic fungal pathogen endemic to Southeast Asia. In the host T. marneffei resides within macrophages where it grows as a fission yeast. T. marneffei has genes of both iron assimilation systems as well as a paralogue of the siderophore biosynthetic gene sidA, designated sidX. Unlike other fungi, deletion of sidA or sidX resulted in cell type-specific effects. Mutant analysis showed that T. marneffei yeast cells also employ RIA for iron acquisition, providing an additional system in this cell type that differs substantially from hyphal cells. These data illustrate the specialized iron acquisition systems used by the different cell types of a dimorphic fungal pathogen and highlight the complexity in siderophore-biosynthetic pathways amongst fungi.

Iron Homeostasis in Mycobacterium tuberculosis: Mechanistic Insights into Siderophore-Mediated Iron Uptake.

Mycobacterium tuberculosis requires iron for normal growth but faces a limitation of the metal ion due to its low solubility at biological pH and the withholding of iron by the mammalian host. The pathogen expresses the Fe(3+)-specific siderophores mycobactin and carboxymycobactin to chelate the metal ion from insoluble iron and the host proteins transferrin, lactoferrin, and ferritin. Siderophore-mediated iron uptake is essential for the survival of M. tuberculosis, as knockout mutants, which were defective in siderophore synthesis or uptake, failed to survive in low-iron medium and inside macrophages. But as excess iron is toxic due to its catalytic role in the generation of free radicals, regulation of iron uptake is necessary to maintain optimal levels of intracellular iron. The focus of this review is to present a comprehensive overview of iron homeostasis in M. tuberculosis that is discussed in the context of mycobactin biosynthesis, transport of iron across the mycobacterial cell envelope, and storage of excess iron. The clinical significance of the serum iron status and the expression of the iron-regulated protein HupB in tuberculosis (TB) patients is presented here, highlighting the potential of HupB as a marker, notably in extrapulmonary TB cases.

Pharmacodynamic Profiling of a Siderophore-Conjugated Monocarbam in Pseudomonas aeruginosa: Assessing the Risk for Resistance and Attenuated Efficacy.

The objective of this study was to investigate the risk of attenuated efficacy due to adaptive resistance for the siderophore-conjugated monocarbam SMC-3176 in Pseudomonas aeruginosa by using a pharmacokinetic/pharmacodynamic (PK/PD) approach. MICs were determined in cation-adjusted Mueller-Hinton broth (MHB) and in Chelex-treated, dialyzed MHB (CDMHB). Spontaneous resistance was assessed at 2× to 16× the MIC and the resulting mutants sequenced. Efficacy was evaluated in a neutropenic mouse thigh model at 3.13 to 400 mg/kg of body weight every 3 h for 24 h and analyzed for association with free time above the MIC (fT>MIC). To closer emulate the conditions of the in vivo model, we developed a novel assay testing activity mouse whole blood (WB). All mutations were found in genes related to iron uptake: piuA, piuC, pirR, fecI, and pvdS. Against four P. aeruginosa isolates, SMC-3176 displayed predictable efficacy corresponding to the fT>MIC using the MIC in CDMHB (R(2) = 0.968 to 0.985), with stasis to 2-log kill achieved at 59.4 to 81.1%. Efficacy did not translate for P. aeruginosa isolate JJ 4-36, as the in vivo responses were inconsistent with fT>MIC exposures and implied a threshold concentration that was greater than the MIC. The results of the mouse WB assay indicated that efficacy was not predictable using the MIC for JJ 4-36 and four additional isolates, against which in vivo failures of another siderophore-conjugated β-lactam were previously reported. SMC-3176 carries a risk of attenuated efficacy in P. aeruginosa due to rapid adaptive resistance preventing entry via the siderophore-mediated iron uptake systems. Substantial in vivo testing is warranted for compounds using the siderophore approach to thoroughly screen for this in vitro-in vivo disconnect in P. aeruginosa.

The N-acetyltransferase gene-implicated iron acquisition contributes to host specificity of Pseudomonas cichorii strain SPC9018 and its virulence

A pathogenicity island within the genome of a multi-host plant bacterium, Pseudomonas cichorii strain SPC9018, comprises the hrp genes encoding a type III secretion system and the pat gene encoding an N-acetyltransferase proposed to play a role in virulence. However, the function of the N-acetyltransferase remains poorly characterized. Interestingly, limiting the iron condition using a phytosiderophore, mugineic acid, resulted in reduced virulence of strain SPC9018 on respective host plants, including eggplant, similar to the reduced virulence observed with a pat gene-deletion mutant. Spectroscopic analyses showed that the pat deletion reduced the concentration of pyoverdine, which is the main siderophore produced by strain SPC9018, leading to a reduction in pyoverdine-mediated iron acquisition. Furthermore, the pat gene deletion mutant showed enhanced expression of the fecA, pvdL, and pvdR genes, whose expression is induced under deficient siderophore-mediated iron uptake. The pat-deletion mutant showed a hyper-swarming phenotype, and the addition of iron decreased this swarming motility. The pat deletion also reduced the adhesion ability of the bacteria, similar to the effect of iron-limited conditions. Furthermore, deletion of the pat gene enhanced expression of the hrp genes. These findings suggested that the pat gene encoding the N-acetyltransferase may be implicated in iron acquisition, contributing to host specificity of P. cichorii strain SPC9018 and its virulence.

Intracellular siderophore but not extracellular siderophore is required for full virulence in Metarhizium robertsii

Efficient iron acquisition mechanisms are fundamental for microbial survival in the environment and for pathogen virulence within their hosts. M. robertsii produces two known iron-binding natural products: metachelins, which are used to scavenge extracellular iron, and ferricrocin, which is strictly intracellular. To study the contribution of siderophore-mediated iron uptake and storage to M. robertsii fitness, we generated null mutants for each siderophore synthase gene (mrsidD and mrsidC, respectively), as well as for the iron uptake transcriptional repressor mrsreA. All of these mutants showed impaired germination speed, differential sensitivity to hydrogen peroxide, and differential ability to overcome iron chelation on growth-limiting iron concentrations. RT-qPCR data supported regulation of mrsreA, mrsidC, and mrsidD by supplied iron in vitro and during growth within the insect host, Spodoptera exigua. We also observed strong upregulation of the insect iron-binding proteins, transferrins, during infection. Insect bioassays revealed that ferricrocin is required for full virulence against S. exigua; neither the loss of metachelin production nor the deletion of the transcription factor mrsreA significantly affected M. robertsii virulence.

Iron acquisition and oxidative stress response in aspergillus fumigatus.

BackgroundAspergillus fumigatus is a ubiquitous airborne fungal pathogen that presents a life-threatening health risk to individuals with weakened immune systems. A. fumigatus pathogenicity depends on its ability to acquire iron from the host and to resist host-generated oxidative stress. Gaining a deeper understanding of the molecular mechanisms governing A. fumigatus iron acquisition and oxidative stress response may ultimately help to improve the diagnosis and treatment of invasive aspergillus infections.ResultsThis study follows a systems biology approach to investigate how adaptive behaviors emerge from molecular interactions underlying A. fumigatus iron regulation and oxidative stress response. We construct a Boolean network model from known interactions and simulate how changes in environmental iron and superoxide levels affect network dynamics. We propose rules for linking long term model behavior to qualitative estimates of cell growth and cell death. These rules are used to predict phenotypes of gene deletion strains. The model is validated on the basis of its ability to reproduce literature data not used in model generation.ConclusionsThe model reproduces gene expression patterns in experimental time course data when A. fumigatus is switched from a low iron to a high iron environment. In addition, the model is able to accurately represent the phenotypes of many knockout strains under varying iron and superoxide conditions. Model simulations support the hypothesis that intracellular iron regulates A. fumigatus transcription factors, SreA and HapX, by a post-translational, rather than transcriptional, mechanism. Finally, the model predicts that blocking siderophore-mediated iron uptake reduces resistance to oxidative stress. This indicates that combined targeting of siderophore-mediated iron uptake and the oxidative stress response network may act synergistically to increase fungal cell killing.Electronic supplementary materialThe online version of this article (doi:10.1186/s12918-015-0163-1) contains supplementary material, which is available to authorized users.

Functional Exposed Amino Acids of BauA as Potential Immunogen Against Acinetobacter baumannii.

Multidrug-resistant Acinetobacter baumannii is recognized to be among the most difficult antimicrobial-resistant gram negative bacilli to control and treat. One of the major challenges that the pathogenic bacteria face in their host is the scarcity of freely available iron. To survive under such conditions, bacteria express new proteins on their outer membrane and also secrete iron chelators called siderophores. Antibodies directed against these proteins associated with iron uptake exert a bacteriostatic or bactericidal effect against A. baumanii in vitro, by blocking siderophore mediated iron uptake pathways. Attempts should be made to discover peptides that could mimic protein epitopes and possess the same immunogenicity as the whole protein. Subsequently, theoretical methods for epitope prediction have been developed leading to synthesis of such peptides that are important for development of immunodiagnostic tests and vaccines. The present study was designed to in silico resolving the major obstacles in the control or in prevention of the diseases caused by A. baumannii. We exploited bioinformatic tools to better understand and characterize the Baumannii acinetobactin utilization structure of A. baumannii and select appropriate regions as effective B cell epitopes. In conclusion, amino acids 26-191 of cork domain and 321-635 of part of the barrel domain including L4-L9, were selected as vaccine candidates. These two regions contain functional exposed amino acids with higher score of B cell epitopes properties. Majority of amino acids are hydrophilic, flexible, accessible, and favorable for B cells from secondary structure point of view.

Plasmid-Encoded Iron Uptake Systems.

Plasmids confer genetic information that benefits the bacterial cells containing them. In pathogenic bacteria, plasmids often harbor virulence determinants that enhance the pathogenicity of the bacterium. The ability to acquire iron in environments where it is limited, for instance the eukaryotic host, is a critical factor for bacterial growth. To acquire iron, bacteria have evolved specific iron uptake mechanisms. These systems are often chromosomally encoded, while those that are plasmid-encoded are rare. Two main plasmid types, ColV and pJM1, have been shown to harbor determinants that increase virulence by providing the cell with essential iron for growth. It is clear that these two plasmid groups evolved independently from each other since they do not share similarities either in the plasmid backbones or in the iron uptake systems they harbor. The siderophores aerobactin and salmochelin that are found on ColV plasmids fall in the hydroxamate and catechol group, respectively, whereas both functional groups are present in the anguibactin siderophore, the only iron uptake system found on pJM1-type plasmids. Besides siderophore-mediated iron uptake, ColV plasmids carry additional genes involved in iron metabolism. These systems include ABC transporters, hemolysins, and a hemoglobin protease. ColV- and pJM1-like plasmids have been shown to confer virulence to their bacterial host, and this trait can be completely ascribed to their encoded iron uptake systems.

Cell growth and protein expression of Shewanella oneidensis in biofilms and hydrogel-entrapped cultures

The performance of biofilm-based bioprocesses is difficult to predict and control because of the intrinsic heterogeneous and dynamic properties of microbial biofilms. Biofilm mimics, such as microbial cells entrapped in polymeric scaffolds that are permeable for nutrients, have been proposed to replace real biofilms to achieve long-term robust performance in engineering applications. However, the physiological differences between cells that are physically entrapped in a synthetic polymeric matrix and biofilm cells that are encased in a self-produced polymeric matrix remain unknown. In this study, using Shewanella oneidensis as a model organism and alginate hydrogel as a model synthetic matrix, we compared the cell growth and protein expression in entrapped cultures and biofilms. The hydrogel-entrapped cultures were found to exhibit a growth rate comparable with biofilms. There was no substantial difference in cell viability, surface charge, as well as hydrophobicity between the cells grown in alginate hydrogel and those grown in biofilms. However, the gel-entrapped cultures were found to be physiologically different from biofilms. The gel-entrapped cultures had a higher demand for metabolic energy. The siderophore-mediated iron uptake was repressed in the gel-entrapped cells. The presence of the hydrogel matrix decreased the expression of proteins involved in biofilm formation, while inducing the production of extracellular DNA (eDNA) in the gel-entrapped cultures. These results advance the fundamental understanding of the physiology of hydrogel-entrapped cells, which can lead to more efficient biofilm mimic-based applications.

The FupA/B protein uniquely facilitates transport of ferrous iron and siderophore-associated ferric iron across the outer membrane of Francisella tularensis live vaccine strain

Francisella tularensis is a highly infectious Gram-negative pathogen that replicates intracellularly within the mammalian host. One of the factors associated with virulence of F. tularensis is the protein FupA that mediates high-affinity transport of ferrous iron across the outer membrane. Together with its paralogue FslE, a siderophore-ferric iron transporter, FupA supports survival of the pathogen in the host by providing access to the essential nutrient iron. The FupA orthologue in the attenuated live vaccine strain (LVS) is encoded by the hybrid gene fupA/B, the product of an intergenic recombination event that significantly contributes to attenuation of the strain. We used (55)Fe transport assays with mutant strains complemented with the different paralogues to show that the FupA/B protein of LVS retains the capacity for high-affinity transport of ferrous iron, albeit less efficiently than FupA of virulent strain Schu S4. (55)Fe transport assays using purified siderophore and siderophore-dependent growth assays on iron-limiting agar confirmed previous findings that FupA/B also contributes to siderophore-mediated ferric iron uptake. These assays further demonstrated that the LVS FslE protein is a weaker siderophore-ferric iron transporter than the orthologue from Schu S4, and may be a result of the sequence variation between the two proteins. Our results indicate that iron-uptake mechanisms in LVS differ from those in Schu S4 and that functional differences in the outer membrane iron transporters have distinct effects on growth under iron limitation.

Identification of an iron acquisition machinery in Flavobacterium columnare

Flavobacterium columnare, a fastidious Gram-negative pathogen and the causative agent of columnaris disease, is one of the most harmful pathogens in the freshwater fish-farming industry. Nevertheless the virulence mechanisms of F. columnare are not well understood. Bacterial iron uptake from the host during infection is an important mechanism of virulence. Here we identified and analyzed part of the iron uptake machinery of F. columnare. Under iron-limited conditions during in vitro growth, synthesis of an outer membrane protein of ~86 kDa was upregulated. This protein was identified as a TonB-dependent ferrichrome-iron receptor precursor (FhuA). Synthesis of siderophores in F. columnare was corroborated by chrome azurol S assays. A putative ferric uptake regulator (Fur) protein was also identified in the F. columnare genome. Structural analysis of the F. columnare Fur protein revealed that it was similar to Fur proteins involved in iron uptake regulation of other bacteria. Furthermore, Salmonella enterica serovar Typhimurium (S. Typhimurium) Δfur mutants were partially complemented by the F. columnare fur gene. We conclude that a siderophore-mediated iron uptake system exists in F. columnare, and fur from F. columnare could partially complement S. Typhimurium Δfur mutant.

An anti-Aspergillusprotein fromEscherichia coliDH5α: Putative inhibitor of siderophore biosynthesis inAspergillus fumigatus

An antifungal protein designated as anti-Aspergillus protein (AAP), produced by Escherichia coli DH5α, was purified and characterised. It exhibited a molecular weight of 60kDa on Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis and depicted 99% purity on ultra performance liquid chromatography. The purified protein manifested antimycotic potential against pathogenic isolates of Aspergillus spp., depicting a minimum inhibitory concentration in the range 15.62-31.25μgml(-1) and 5.0-10.0μg per disc, using microbroth dilution, spore germination inhibition and disc diffusion assays respectively. In vitro toxicity tests demonstrated that it showed no toxicity against human erythrocytes at doses up to 1000μgml(-1) . Matrix-assisted laser desorption ionisation-Time-of-flight analysis of trypsin-digested peptides of purified protein and subsequent Mascot search revealed that several peptides of AAP have identity with bacterial siderophore biosynthetic protein, i.e. non-ribosomal peptide synthetase enzyme, involved in critical step of fungal siderophore biosynthesis. Siderophore-based inhibition was further corroborated by Chrome azurol S assay. Hence, the antagonistic effect might be the result of impediment in siderophore-mediated iron uptake and transport process which may cause critical consequences on Aspergillus growth and virulence.

Pseudomonas aeruginosa Uses Multiple Pathways To Acquire Iron during Chronic Infection in Cystic Fibrosis Lungs

Pseudomonas aeruginosa chronically infects the lungs of more than 80% of adult patients with cystic fibrosis (CF) and is a major contributor to the progression of disease pathology. P. aeruginosa requires iron for growth and has multiple iron uptake systems that have been studied in bacteria grown in laboratory culture. The purpose of this research was to determine which of these are active during infection in CF. RNA was extracted from 149 sputum samples obtained from 23 CF patients. Reverse transcription-quantitative real-time PCR (RT-qPCR) was used to measure the expression of P. aeruginosa genes encoding transport systems for the siderophores pyoverdine and pyochelin, for heme, and for ferrous ions. Expression of P. aeruginosa genes could be quantified in 89% of the sputum samples. Expression of genes associated with siderophore-mediated iron uptake was detected in most samples but was at low levels in some samples, indicating that other iron uptake mechanisms are active. Expression of genes encoding heme transport systems was also detected in most samples, indicating that heme uptake occurs during infection in CF. feoB expression was detected in all sputum samples, implying an important role for ferrous ion uptake by P. aeruginosa in CF. Our data show that multiple P. aeruginosa iron uptake mechanisms are active in chronic CF infection and that RT-qPCR of RNA extracted from sputum provides a powerful tool for investigating bacterial physiology during infection in CF.

A nonribosomal peptide synthetase mediates siderophore production and virulence in the citrus fungal pathogen Alternaria alternata.

Alternaria species produce and excrete dimethyl coprogen siderophores to acquire iron. The Alternaria alternata gene AaNPS6, encoding a polypeptide analogous to fungal nonribosomal peptide synthetases, was found to be required for the production of siderophores and virulence on citrus. Siderophores purified from culture filtrates of the wild-type strain did not induce any phytotoxicity on the leaves of citrus. Fungal strains lacking AaNPS6 produced little or no detectable extracellular siderophores and displayed an increased sensitivity to H₂O₂, superoxide-generating compounds (KO₂ and menadione) and iron depletion. Δnps6 mutants were also defective for the production of melanin and conidia. The introduction of a wild-type AaNPS6 under the control of its endogenous promoter to a Δnps6 null mutant at least partially restored siderophore production and virulence to citrus, demonstrating a functional link between iron uptake and fungal pathogenesis. Elevated sensitivity to H₂O₂, seen for the Δnps6 null strain could be relieved by exogenous application of ferric iron. The expression of the AaNPS6 gene was highly up-regulated under low-iron conditions and apparently controlled by the redox-responsive yeast transcriptional regulator YAP1. Hence, the maintenance of iron homeostasis via siderophore-mediated iron uptake also plays an important role in resistance to toxic reactive oxygen species (ROS). Our results demonstrate further the critical role of ROS detoxification for the pathogenicity of A. alternata in citrus.

Effect of the amino acid substitution in the DNA-binding domain of the Fur regulator on production of pyoverdine

The ferric uptake regulator gene (fur), its promoter region and Fur box of pvdS gene involved in siderophore-mediated iron uptake system were sequenced in the parent strain Pseudomonas aeruginosa PAO1 and in the fur mutant FPA121 derived from the strain PAO1. We identified the gene fur 179 bearing a novel, single-point mutation that changed the amino acid residue Gln60Pro in the DNA-binding domain of the Fur protein. The synthesis of pyoverdine was studied in cultures of the strains PAO1 and FPA121 grown in iron-deplete and iron-replete (60μmol/L FeIII) medium. The amino acid replacement in the regulatory Fur protein is responsible for the overproduction of pyoverdine in iron-deplete and iron-replete medium. No mutation was identified in the Fur box of the gene pvdS.

Siderophore-Mediated Iron Uptake Promotes Yeast–Bacterial Symbiosis

In the present study, siderophore produced by the marine yeast Aureobasidium pullulans was characterized as hydroxamate by chemical and bioassays. The hydroxamate assignment was supported by the appearance of peaks at 1,647.21-1,625.99cm(-1) and at 1,435.04cm(-1) in the infrared spectrum. The purified siderophore exhibited specific growth-promoting activity under iron-limited conditions for siderophore auxotrophic probiotic bacteria. Cross-utilization of siderophore indicates a symbiotic relationship between the yeast A. pullulans and the selected probiotic bacterial strains. Statistical optimization of medium components for improved siderophore production in A. pullulans was depicted by response surface methodology. The shift in UV-Vis spectroscopy indicates the photoreactive property and subsequent oxidative cleavage of purified siderophore on exposure to sunlight.