Humic acid and iron uptake by plants

Summary The behaviour of ferric EDTA and ferric citrate in nutrient solution and their interaction with humic acid was investigated at various hydrogen ion concentrations using the technique of membrane ultrafiltration to separate small iron species from high molecular weight products of hydrolysis and to estimate the binding of iron by humic acid. Ferric EDTA was found to be of small molecular size at all pH values between 5.0 and 7.0 whilst ferric citrate solutions contained an increasing proportion of high molecular weight material as pH was increased from 5.0 to 7.0. Some iron present in solutions of both ferric EDTA and ferric citrate was bound by humic acid at all pH values from 5.0 to 7.0. Studies were also made of the uptake of iron by wheat roots from nutrient solutions containing either ferric EDTA or ferric citrate and of the effect of humic acid on uptake. More iron was absorbed from ferric EDTA than from ferric citrate at all pH values. Increasing pH between 5.0 and 7.0 resulted in a progressive decrease in the uptake of iron in both cases. The presence of humic acid depressed iron absorption from both solutions at all pH values.     

Effects of iron sources on the growth and lipid/carbohydrate production of marine microalga <Emphasis Type="Italic">Dunaliella tertiolecta</Emphasis>

The effects of iron sources with different speciation and anionic moieties (ferric chloride, ferrous chloride, ferric EDTA, ferrous EDTA, ferric ammonium sulfate, and ferrous ammonium sulfate) on the cell growth and the production of energy storage (lipid and carbohydrate) from Dunaliella tertiolecta were investigated. The influence of iron dosage was also compared in the range from 0.65 mg/L (1X) to 6.5 mg/L (10X) as Fe concentration. Best cell growth rate was achieved when ferrous ammonium sulfate was used. Ferric EDTA resulted in higher lipid content than other iron sources, while ferrous ammonium sulfate favored the accumulation of carbohydrate among six iron sources. The accumulations of lipid and carbohydrate as energy storage competed each other and thus both contents did not increase together. In the presence of ferric EDTA, lipid content is increasing, while carbohydrate content is decreasing. On the contrary, lipid content is decreasing while carbohydrate is increasing in the presence of ferric ammonium sulfate. Because the overall carbohydrate content was larger than that of lipid, bioethanol production would be more advantageous than biodiesel production with the present D. tertiolecta strain if the carbohydrate in D. tertiolecta contains a high fraction of glucose with a good saccharification yield.     

The role of reduced iron powder in the fermentative production of tetanus toxin

When tetanus toxin is made by fermentation with Clostridium tetani, the traditional source of iron is an insoluble preparation called reduced iron powder. This material removes oxygen from the system by forming FeO₂ (rust). When inoculated in a newly developed medium lacking animal and dairy products and containing glucose, soy-peptone, and inorganic salts, growth and toxin production were poor without reduced iron powder. The optimum concentration of reduced iron powder for toxin production was found to be 0.5 g/l. Growth was further increased by higher concentrations, but toxin production decreased. Inorganic iron sources failed to replace reduced iron powder for growth or toxin formation. The iron source that came closest was ferrous ammonium sulfate. The organic iron sources ferric citrate and ferrous gluconate were more active than the inorganic compounds but could not replace reduced iron powder. Insoluble iron sources, such as iron wire, iron foil, and activated charcoal, were surprisingly active. Combinations of activated charcoal with soluble iron sources such as ferrous sulfate, ferric citrate, and ferrous gluconate showed increased activity, and the ferrous gluconate combination almost replaced reduced iron powder. It thus appears that the traditional iron source, reduced iron powder, plays a double role in supporting tetanus toxin formation, i.e., releasing soluble sources of iron and providing an insoluble surface.     

Mechanisms of ferric and ferrous iron uptake by Bifidobacterium bifidum var. pennsylvanicus

Iron uptake studies in Bifidobacterium bifidum var. pennsylvanicus were carried out using ferric citrate at iron concentrations above 0.01 mM and pH 7, ferrous iron at concentrations less than 0.01 mM at pH 5. Two ferric iron transport systems were distinguished: the temperature-insensitive polymer, and the temperature-sensitive monomer uptake. Both showed a saturation phenomenon. The transport of ferrous iron at concentrations below 0.01 mM was temperature-dependent, and its affinity for iron was higher than that of a system operating at iron concentrations higher than 0.01 mM. The use of various metabolic inhibitors indicated that ferrous iron transport at pH 5 at both high and low iron concentrations was mediated by transport-type ATPase. Proton gradient dissipators abolished ferrous iron uptakes as well as the ferric monomer uptake. Uptake of the ferric polymer was insensitive to metabolic inhibitors. The functional significance of the various types of iron transport systems may be related to the nutritional immunity phenomenon.     

Transport and utilization of ferrioxamine-E-bound iron inErwinia herbicola (Pantoea agglomerans)

Summary We have analyzed ferrioxamine-E-mediated iron uptake and metabolization inErwinia herbicola K4 (Pantoea agglomerans) by means of in vivo Mössbauer spectroscopy and radioactive labeling techniques. A comparison of cell spectra with the spectrum of ferrioxamine clearly demonstrates that ferrioxamine E is not accumulated in the cell, indicating a fast metal transfer. Only two major components of iron metabolism can be detected, a ferric and a ferrous species. At 30 min after uptake, 86% of the internalized metal corresponded to a ferrous ion compound and 14% to a ferric iron species. Metal transfer apparently involves a reductive process. With progressing growth, the oxidized species of the two major proteins becomes dominant. The two iron metabolites closely resemble species previously isolated fromEscherichia coli. These components of iron metabolism differ from bacterio-ferritin, cytochromes and most iron-sulfur proteins. All other iron-containing cellular components are at least one order of magnitude lower in concentration. We suggest that the ferrous and ferric iron species correspond to two different oxidation states of a low-molecular mass protein.     

Toxicity and bioaccumulation of iron in soil microalgae

Microalgae are extensively used in the remediation of heavy metals like iron. However, factors like toxicity, bioavailability and iron speciation play a major role in its removal by microalgae. Thus, in this study, toxicity of three different iron salts (FeSO₄, FeCl₃ and Fe(NO₃)₃) was evaluated towards three soil microalgal isolates, Chlorella sp. MM3, Chlamydomonas sp. MM7 and Chlorococcum sp. MM11. Interestingly, all the three iron salts gave different EC50 concentrations; however, ferric nitrate was found to be significantly more toxic followed by ferrous sulphate and ferric chloride. The EC₅₀ analysis revealed that Chlorella sp. was significantly resistant to iron compared to other microalgae. However, almost 900 μg g^?1 iron was accumulated by Chlamydomonas sp. grown with 12 mg L^?1 ferric nitrate as an iron source when compared to other algae and iron salts. The time-course bioaccumulation confirmed that all the three microalgae adsorb the ferric salts such as ferric nitrate and ferric chloride more rapidly than ferrous salt, whereas intracellular accumulation was found to be rapid for ferrous salts. However, the amount of iron accumulated or adsorbed by algae, irrespective of species, from ferrous sulphate medium is comparatively lower than ferric chloride and ferric nitrate medium. The Fourier transform infrared spectroscopy (FTIR) analysis shows that the oxygen atom and P?=?O group of polysaccharides present in the cell wall of algae played a major role in the bioaccumulation of iron ions by algae.     

Evaluation of different iron compounds in chlorotic Italian lemon trees (Citrus lemon).

The severe deficiency of iron or ferric chlorosis is a serious problem of most citrus trees established in calcareous soils, as a result of the low availability of iron in these soils and the poor uptake and limited transport of this nutrient in trees. The objective of this study was to evaluate the response of chlorotic Italian lemon trees (Citrus lemon) to the application of iron compounds to roots and stems. On comparing the effects of aqueous solutions of ferric citrate, ferrous sulphate and FeEDDHA chelate, applied to 20% of the roots grown in soil and sand, of trees that were planted in pots containing calcareous soil, it was observed that the chelate fully corrected ferric chlorosis, while citrate and sulphate did not solve the problem. EDDHA induced the root uptake of iron as well as the movement of the nutrient up to the leaves. With the use of injections of ferric solutions into the secondary stem of adult trees, ferric citrate corrected chlorosis but ferrous sulphate did not. The citrate ion expanded the mobility of iron within the plant, from the injection points up to the leaves, whereas the sulphate ion did not sufficiently improve the movement of iron towards the leaf mesophyll.     

Ascorbate Efflux as a New Strategy for Iron Reduction and Transport in Plants

Iron (Fe) is essential for virtually all living organisms. The identification of the chemical forms of iron (the speciation) circulating in and between cells is crucial to further understand the mechanisms of iron delivery to its final targets. Here we analyzed how iron is transported to the seeds by the chemical identification of iron complexes that are delivered to embryos, followed by the biochemical characterization of the transport of these complexes by the embryo, using the pea (Pisum sativum) as a model species. We have found that iron circulates as ferric complexes with citrate and malate (Fe(III)₃Cit₂Mal₂, Fe(III)₃Cit₃Mal₁, Fe(III)Cit₂). Because dicotyledonous plants only transport ferrous iron, we checked whether embryos were capable of reducing iron of these complexes. Indeed, embryos did express a constitutively high ferric reduction activity. Surprisingly, iron(III) reduction is not catalyzed by the expected membrane-bound ferric reductase. Instead, embryos efflux high amounts of ascorbate that chemically reduce iron(III) from citrate-malate complexes. In vitro transport experiments on isolated embryos using radiolabeled ⁵⁵Fe demonstrated that this ascorbate-mediated reduction is an obligatory step for the uptake of iron(II). Moreover, the ascorbate efflux activity was also measured in Arabidopsis embryos, suggesting that this new iron transport system may be generic to dicotyledonous plants. Finally, in embryos of the ascorbate-deficient mutants vtc2-4, vtc5-1, and vtc5-2, the reducing activity and the iron concentration were reduced significantly. Taken together, our results identified a new iron transport mechanism in plants that could play a major role to control iron loading in seeds.     

Lipid peroxidation of rabbit small intestinal microvillus membrane vesicles by iron complexes

Fe(II)- and Fe(III)-induced lipid peroxidation of rabbit small intestinal microvillus membrane vesicles was studied. Ferrous ammonium sulphate, ferrous ascorbate at a molar ratio of 10:1, and ferric citrate, at molar ratios of 1:1 and 1:20, did not stimulate lipid peroxidation. Ferrous ascorbate, 1:1, induced low stimulation, while ferrous ascorbate, 1:20 gave higher stimulation of lipid peroxidation. These results show that in our experimental system, ascorbate is a promotor rather than an inhibitor of lipid peroxidation. Ferric nitrilotriacetate (at molar ratios of 1:2 and 1:10), at an iron concentration of 200 microM, was by far the most effective in inducing lipid peroxidation. Superoxide dismutase, mannitol and glutathione had no effect, while catalase, thiourea and vitamin E markedly decreased ferrous ascorbate 1:20-induced lipid peroxidation. Ferric nitrilotriacetate-induced lipid peroxidation was slightly reduced by catalase and mannitol, significantly reduced by superoxide dismutase, and completely inhibited by thiourea. Glutathione caused a 100% increase in the ferric nitrilotriacetate-induced lipid peroxidation. These results suggest that Fe(II) in the presence of trace amounts of Fe(III), or an oxidizing agent and Fe(III) in the presence of Fe(II) or a reducing agent, are potent stimulators of lipid peroxidation of microvillus membrane vesicles. Addition of deferoxamine completely inhibited both ferrous ascorbate, 1:20 and ferric nitrilotriacetate-induced lipid peroxidation, demonstrating the requirement for iron for its stimulation. Iron-induced peroxidation of microvillus membrane may have physiological significance because it could already be demonstrated at 2 microM iron concentration.     

Non-transferrin iron reduction and uptake are regulated by transmembrane ascorbate cycling in K562 cells

K562 erythroleukemia cells import non-transferrin-bound iron (NTBI) by an incompletely understood process that requires initial iron reduction. The mechanism of NTBI ferrireduction remains unknown but probably involves transplasma membrane electron transport. We here provide evidence for a novel mechanism of NTBI reduction and uptake by K562 cells that utilizes transplasma membrane ascorbate cycling. Incubation of cells with dehydroascorbic acid, but not ascorbate, resulted in (i) accumulation of intracellular ascorbate that was blocked by the glucose transporter inhibitor, cytochalasin B, and (ii) subsequent release of micromolar concentrations of ascorbate into the external medium via a route that was sensitive to the anion channel inhibitor, 4,4'-diisothiocyanatostilbene-2,2'-disulfonate. Ascorbate-deficient control cells demonstrated low levels of ferric citrate reduction. However, incubation of the cells with dehydroascorbic acid resulted in a dose-dependent stimulation of both iron reduction and uptake from radiolabeled [(55)Fe]ferric citrate. This stimulation was abrogated by ascorbate oxidase treatment, suggesting dependence on direct chemical reduction by ascorbate. These results support a novel model of NTBI reduction and uptake by K562 cells in which uptake is preceded by reduction of iron by extracellular ascorbate, the latter of which is subsequently regenerated by transplasma membrane ascorbate cycling.     

Vibrio cholerae VciB promotes iron uptake via ferrous iron transporters

Vibrio cholerae uses a variety of strategies for obtaining iron in its diverse environments. In this study we report the identification of a novel iron utilization protein in V. cholerae, VciB. The vciB gene and its linked gene, vciA, were isolated in a screen for V. cholerae genes that permitted growth of an Escherichia coli siderophore mutant in low-iron medium. The vciAB operon encodes a predicted TonB-dependent outer membrane receptor, VciA, and a putative inner membrane protein, VciB. VciB, but not VciA, was required for growth stimulation of E. coli and Shigella flexneri strains in low-iron medium. Consistent with these findings, TonB was not needed for VciB-mediated growth. No growth enhancement was seen when vciB was expressed in an E. coli or S. flexneri strain defective for the ferrous iron transporter Feo. Supplying the E. coli feo mutant with a plasmid encoding either E. coli or V. cholerae Feo, or the S. flexneri ferrous iron transport system Sit, restored VciB-mediated growth; however, no stimulation was seen when either of the ferric uptake systems V. cholerae Fbp and Haemophilus influenzae Hit was expressed. These data indicate that VciB functions by promoting iron uptake via a ferrous, but not ferric, iron transport system. VciB-dependent iron accumulation via Feo was demonstrated directly in iron transport assays using radiolabeled iron. A V. cholerae vciB mutant did not exhibit any growth defects in either in vitro or in vivo assays, possibly due to the presence of other systems with overlapping functions in this pathogen.     

Uptake of ionic 55Fe by mouse peritoneal macrophages in vitro.

Mouse peritoneal macrophages were maintained in vitro up to 3 days and exposed to radiolabelled ⁵⁵Fe in the form of ferrous citrate, ferrous sulfate, and ferric chloride in concentrations of 3–5 γ Fe/ml. The divalent iron compounds were taken up 10–40 times more extensively per weight of iron than the trivalent iron compounds. The net uptake of ferrous citrate was linear during the first day and thereafter increased at a slower rate. Macrophages in culture for 1 week showed one-third the average uptake of freshly cultured cells during comparable periods of exposure to ferrous citrate. The iron taken up was used in the synthesis of mouse ferritin. Uptake of ferrous citrate was influenced by serum concentration in the tissue culture medium, temperature, pinocytosis and phagocytosis of both latex particles and heated rat erythrocytes. Uptake of ferrous citrate was enhanced by exposure to either sodium fluoride (5×10^?3 M), or 2,4-dinitrophenol (1×10^?5 M), but was not affected by cyanide, azide, or cycloheximide. The effect of sodium fluoride was not demonstrated when ferrous sulfate was substituted for ferrous citrate. The results reported here suggest that the ability of macrophages to take up ferrous citrate is good in freshly explanted cultures, is a temperature-dependent process, is suppressed by pinocytosis and phagocytosis, and paradoxically enhanced by certain metabolic inhibitors.     

Separate pathways for cellular uptake of ferric and ferrous iron

Separate pathways for transport of nontransferrin ferric and ferrous iron into tissue cultured cells were demonstrated. Neither the ferric nor ferrous pathway was shared with either zinc or copper. Manganese shared the ferrous pathway but had no effect on cellular uptake of ferric iron. We postulate that ferric iron was transported into cells via beta(3)-integrin and mobilferrin (IMP), whereas ferrous iron uptake was facilitated by divalent metal transporter-1 (DMT-1; Nramp-2). These conclusions were documented by competitive inhibition studies, utilization of a beta(3)-integrin antibody that blocked uptake of ferric but not ferrous iron, development of an anti-DMT-1 antibody that blocked ferrous iron and manganese uptake but not ferric iron, transfection of DMT-1 DNA into tissue culture cells that showed enhanced uptake of ferrous iron and manganese but neither ferric iron nor zinc, hepatic metal concentrations in mk mice showing decreased iron and manganese but not zinc or copper, and data showing that the addition of reducing agents to tissue culture media altered iron binding to proteins of the IMP and DMT-1 pathways. Although these experiments show ferric and ferrous iron can enter cells via different pathways, they do not indicate which pathway is dominant in humans.     

Genetics and environmental regulation of Shigella iron transport systems

Shigella spp. have transport systems for both ferric and ferrous iron. The iron can be taken up as free iron or complexed to a variety of carriers. All Shigella species have both the Feo and Sit systems for acquisition of ferrous iron, and all have at least one siderophore-mediated system for transport of ferric iron. Several of the transport systems, including Sit, Iuc/IutA (aerobactin synthesis and transport), Fec (ferric di-citrate uptake), and Shu (heme transport) are encoded within pathogenicity islands. The presence and the genomic locations of these islands vary considerably among the Shigella species, and even between isolates of the same species. The expression of the iron transport systems is influenced by the concentration of iron and by environmental conditions including the level of oxygen. ArcA and FNR regulate iron transport gene expression as a function of oxygen tension, with the sit and iuc promoters being highly expressed in aerobic conditions, while the feo ferrous iron transporter promoter is most active under anaerobic conditions. The effects of oxygen are also seen in infection of cultured cells by Shigella flexneri; the Sit and Iuc systems support plaque formation under aerobic conditions, whereas Feo allows plaque formation anaerobically.     

Effects of high stability iron-complexes on the kinetics of iron accumulation and excretion in Mytilus edvlis (L.)

The effects of small molecular weight complexes with a high affinity for iron on the uptake, accumulation and excretion of iron by the common mussel, Mytilus edulis (L.) have been investigated. Fe(III) complexes of citrate, EDTA and 1,10-phenanthroline increased both the rate of uptake and the total amount accumulated when compared to those for particulate ferric hydroxide in sea water, whereas ferrichrome b and the Fe(III) complexes of aceto- and benzo-hydroxamic acids give a decrease. Increased uptake is, however, compensated for by an increased rate of excretion resulting in an almost constant residence time for this metal. The iron is accumulated principally in the viscera with a smaller but significant proportion in the gills. The effects produced by prior complexation of the iron cannot be correlated with either the strength of binding of the complex to the iron or the exchangeability of the iron with other ligands but may be concerned with the endocytosis, the mechanism of uptake for iron previously shown to occur in Mytilus.     

Uptake of ionic Fe by mouse peritoneal macrophages in vitro

Mouse peritoneal macrophages were maintained in vitro up to 3 days and exposed to radiolabelled ⁵⁵Fe in the form of ferrous citrate, ferrous sulfate, and ferric chloride in concentrations of 3–5 γ Fe/ml. The divalent iron compounds were taken up 10–40 times more extensively per weight of iron than the trivalent iron compounds. The net uptake of ferrous citrate was linear during the first day and thereafter increased at a slower rate. Macrophages in culture for 1 week showed one-third the average uptake of freshly cultured cells during comparable periods of exposure to ferrous citrate. The iron taken up was used in the synthesis of mouse ferritin. Uptake of ferrous citrate was influenced by serum concentration in the tissue culture medium, temperature, pinocytosis and phagocytosis of both latex particles and heated rat erythrocytes. Uptake of ferrous citrate was enhanced by exposure to either sodium fluoride (5×10⁻³ M), or 2,4-dinitrophenol (1×10⁻⁵ M), but was not affected by cyanide, azide, or cycloheximide. The effect of sodium fluoride was not demonstrated when ferrous sulfate was substituted for ferrous citrate. The results reported here suggest that the ability of macrophages to take up ferrous citrate is good in freshly explanted cultures, is a temperature-dependent process, is suppressed by pinocytosis and phagocytosis, and paradoxically enhanced by certain metabolic inhibitors.     

Nonreductive Iron Uptake Mechanism in the Marine Alveolate Chromera velia

Chromera velia is a newly cultured photosynthetic marine alveolate. This microalga has a high iron requirement for respiration and photosynthesis, although its natural environment contains less than 1 nm of this metal. We found that this organism uses a novel mechanism of iron uptake, differing from the classic reductive and siderophore-mediated iron uptake systems characterized in the model yeast Saccharomyces cerevisiae and present in most yeasts and terrestrial plants. C. velia has no trans-plasma membrane electron transfer system, and thus cannot reduce extracellular ferric chelates. It is also unable to use hydroxamate siderophores as iron sources. Iron uptake from ferric citrate by C. velia is not inhibited by a ferrous chelator, but the rate of uptake is strongly decreased by increasing the ferric ligand (citrate) concentration. The cell wall contains a large number of iron binding sites, allowing the cells to concentrate iron in the vicinity of the transport sites. We describe a model of iron uptake in which aqueous ferric ions are first concentrated in the cell wall before being taken up by the cells without prior reduction. We discuss our results in relation to the strategies used by the phytoplankton to take up iron in the oceans.Chromera velia is a newly cultured marine alveolate containing a photosynthetic plastid phylogenetically related to vestigial plastids in apicomplexan (Moore et al., 2008). It represents the closest free-living photosynthetic relative to apicomplexan parasites, thus providing a powerful model to study the evolution of eukaryotic adaptability (Moore et al., 2008). To gain further insight into the biology of this organism, the genome of which remains unsequenced, we investigated its iron metabolism and its mechanisms of iron uptake. We compared the data obtained with other phytoplanktonic organisms sharing the same ecological niche, and with a terrestrial unicellular eukaryote, the yeast Saccharomyces cerevisiae. S. cerevisiae is phylogenetically distant from C. velia, but its mechanisms of iron uptake are well characterized, and thus constitutes a useful model in these studies.Iron uptake by terrestrial microorganisms and plants is mostly based on the use of two main strategies, both of which have been previously characterized in S. cerevisiae. The first strategy is the reductive mechanism of uptake. Extracellular ferric complexes are first dissociated by reduction, via trans-plasma membrane electron transfer catalyzed by specialized flavohemoproteins (Fre). Free iron is then imported by a high-affinity permease system (Ftr1) coupled to a copper-dependent oxidase (Fet3), allowing iron to be channeled through the plasma membrane. In the second strategy, the siderophore-mediated mechanism, siderophores excreted by the cells or produced by other bacterial or fungal species are taken up without prior dissociation, via specific, copper-independent high-affinity receptors. Iron is then dissociated from the siderophores inside the cells, probably by reduction (for review, see Kosman, 2003; Philpott, 2006). Chlamydomonas reinhardtii is a photosynthetic eukaryotic model organism for the study of iron homeostasis, which shares with yeast the strategy 1 of iron uptake (copper-dependent reductive iron uptake; Merchant et al., 2006).Much less is known about the strategies used by marine phytoplankton to acquire iron. Some data suggest that these two strategies are used by some marine microalgae (Soria-Dengg and Horstmann, 1995; Allen et al., 2008). However, for most marine unicellular eukaryotes the mechanisms of iron assimilation are completely unknown. The strategies used by these organisms to acquire iron must have evolved to adapt to the very particular conditions that prevail in their surrounding natural environment: The transition metal composition of the ocean differs greatly from that of terrestrial environments (Butler, 1998). In particular, iron levels in surface seawater are extremely low (0.02–1 nm; Turner et al., 2001). Therefore a strategy of iron uptake operating efficiently in a terrestrial environment that contains iron at a micromolar level may be inefficient in a marine environment. No classic iron uptake system with an affinity constant in the nanomolar range has ever been found. Additionally, the marine environment imposes physical limits on the classic strategies of uptake, including the high diffusion rate of the species of interest (siderophores or reduced iron; Völker and Wolf-Gladrow, 1999). It is well known that the low levels of iron limits primary production of phytoplankton and carbon fluxes across vast regions of the world’s oceans (Coale et al., 2004; Pollard et al., 2009). It is thus of particular interest to elucidate the molecular mechanisms underlying acquisition of iron by marine phytoplankton and to determine which iron sources are preferentially assimilated with regards to the yield of carbon fixation.In this study, we investigated the mechanisms of iron uptake by C. velia, and found that this organism uses a nonreductive uptake system of ferric ions, which are first concentrated in the cell wall. Our findings provide a better understanding of the biology of this organism, and highlights the need for further study on the mechanisms of iron acquisition in marine phytoplankton.     

Effects of porphyrins and inorganic iron on the growth of Prevotella intermedia

We demonstrated earlier that hemin-iron-containing compounds which include hemin, human hemoglobin, bovine hemoglobin, and bovine catalase stimulate the growth of Prevotella intermedia [Leung, Subramaniam, Okamoto, Fukushima, Lai, FEMS Microbiol. Lett. 162 (1998) 227-233]. However, the contributions of tetrapyrrole porphyrin ring in these hemin-iron sources as well as inorganic iron for the growth of this organism have not been determined. The purpose of this study was to examine the effects of porphyrins, host iron-binding proteins, and various inorganic iron sources on the growth of hemin-iron depleted P. intermedia. Protoporphyrin IX and protoporphyrin IX-zinc, either in the presence or absence of supplemented ferrous or ferric iron, promoted the growth of P. intermedia at a rate that was comparable to that of the hemin control. On the other hand, neither the host iron proteins, transferrin and lactoferrin, nor the inorganic iron sources which included ferrous chloride, ferric chloride, ferric citrate, ferric nitrate, and ferric ammonium citrate at concentrations up to 200 microM stimulated the growth of hemin-iron-restricted P. intermedia. The results suggest that P. intermedia only use iron in a specific form and that the porphyrin-ring structure is essential for the growth of P. intermedia as in the case of other related organisms.     

Rapid evolution of a bacterial iron acquisition system

Under iron limitation, bacteria scavenge ferric (Fe³⁺) iron bound to siderophores or other chelates from the environment to fulfill their nutritional requirement. In gram‐negative bacteria, the siderophore uptake system prototype consists of an outer membrane transporter, a periplasmic binding protein and a cytoplasmic membrane transporter, each specific for a single ferric siderophore or siderophore family. Here, we show that spontaneous single gain‐of‐function missense mutations in outer membrane transporter genes of Bradyrhizobium japonicum were sufficient to confer on cells the ability to use synthetic or natural iron siderophores, suggesting that selectivity is limited primarily to the outer membrane and can be readily modified. Moreover, growth on natural or synthetic chelators required the cytoplasmic membrane ferrous (Fe²⁺) iron transporter FeoB, suggesting that iron is both dissociated from the chelate and reduced to the ferrous form within the periplasm prior to cytoplasmic entry. The data suggest rapid adaptation to environmental iron by facile mutation of selective outer membrane transporter genes and by non‐selective uptake components that do not require mutation to accommodate new iron sources.