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Cited by 95Antifungal resistance and susceptibilityFungal Infections and StudiesFungal and yeast genetics research

Host Iron Withholding Demands Siderophore Utilization for Candida glabrata to Survive Macrophage Killing

Open access
Nevitt, Tracy, Thiele, Dennis J.·2011·PLoS Pathogens
DOI 10.1371/journal.ppat.1001322PMC PMC3060170
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Abstract

The fungal pathogen Candida glabrata has risen from an innocuous commensal to a major human pathogen that causes life-threatening infections with an associated mortality rate of up to 50%. The dramatic rise in the number of immunocompromised individuals from HIV infection, tuberculosis, and as a result of immunosuppressive regimens in cancer treatment and transplant interventions have created a new and hitherto unchartered niche for the proliferation of C. glabrata. Iron acquisition is a known microbial virulence determinant and human diseases of iron overload have been found to correlate with increased bacterial burden. Given that more than 2 billion people worldwide suffer from iron deficiency and that iron overload is one of the most common single-gene inherited diseases, it is important to understand whether host iron status may influence C. glabrata infectious disease progression. Here we identify Sit1 as the sole siderophore-iron transporter in C. glabrata and demonstrate that siderophore-mediated iron acquisition is critical for enhancing C. glabrata survival to the microbicidal activities of macrophages. Within the Sit1 transporter, we identify a conserved extracellular SIderophore Transporter Domain (SITD) that is critical for siderophore-mediated ability of C. glabrata to resist macrophage killing. Using macrophage models of human iron overload disease, we demonstrate that C. glabrata senses altered iron levels within the phagosomal compartment. Moreover, Sit1 functions as a determinant for C. glabrata to survive macrophage killing in a manner that is dependent on macrophage iron status. These studies suggest that host iron status is a modifier of infectious disease that modulates the dependence on distinct mechanisms of microbial Fe acquisition.

AI snapshot

Written from the paper’s full text.

TL;DR

Candida glabrata encodes a single siderophore-iron transporter, Sit1, and siderophore-mediated iron acquisition via Sit1 enhances its survival to macrophage killing. Macrophage iron status inversely determines C. glabrata dependence on Sit1/siderophore utilization, with iron-loaded macrophages reducing that dependence and iron-restricted macrophages increasing it.

Key findings

  • C. glabrata has one predicted siderophore transporter, Sit1 (CAGL0E04092g), with 72% identity to S. cerevisiae Arn1; sit1Δ cannot use ferrichrome, ferrirubin, or coprogen, while wild-type and SIT1-reconstituted strains grow on these siderophores under iron deficiency.
  • SIT1 mRNA and Sit1 protein accumulate under iron deficiency, and Sit1 localizes to the plasma membrane under iron deficiency even without ferrichrome; siderophore utilization remains functional under copper deficiency imposed by BCS.
  • In J774A.1 mouse and U937 human macrophages, wild-type and SIT1-reconstituted C. glabrata grown with 10 µM ferrichrome showed enhanced survival versus sit1Δ; loading macrophages with 20 µM ferric ammonium citrate for 2 days increased CFU and abolished the siderophore-dependent differences.
  • Macrophage iron status dictated Sit1 dependence: FPNH32R-expressing iron-loaded macrophages showed no siderophore-dependent survival differences, whereas hepcidin-treated FPNC326Y-expressing iron-restricted macrophages imposed greater dependence; SIT1 transcript levels were lower in C. glabrata recovered from FPNH32R versus FPNC326Y macrophages. Primary macrophages from flatiron (ffe) mice also showed diminished siderophore-dependent survival.
  • The conserved Sit1 SITD residue Y575 is required for full ferrichrome utilization and for the siderophore-dependent survival advantage in macrophages; Sit1Y575A localized to the plasma membrane but was impaired in ferrichrome use.

Methods

  • Yeast genetics and growth: C. glabrata BG2/BG14, sit1Δ, SIT1-reconstituted, Sit1Flag/mCherry, and SIT1Y575A strains; iron-replete medium with 5 µM ferrous ammonium sulfate, iron-deficient medium with 100 µM BPS, spot assays with 10 µM siderophores, RNA blot, semi-quantitative RT-PCR, immunoblot, and fluorescence/indirect immunofluorescence microscopy.
  • Macrophage infection: J774A.1 mouse and differentiated U937 human macrophages activated with IFN-gamma/LPS or LPS, with or without 20 µM ferric ammonium citrate for 2 days; infected at MOI 1:4 or 1:1; CFU quantified after macrophage lysis. Primary bone marrow macrophages from control C3H and flatiron (ffe) mice were also infected. Stable J774A.1 lines expressed wild-type FPN, FPNH32R, or FPNC326Y; ferritin immunoblot, TNF-alpha ELISA, and ROS measurement were used.
  • Analysis: one-way ANOVA; results shown as mean survival relative to wild-type C. glabrata grown under iron deficiency; each experiment included 6 replicates per sample and was representative of at least 3 experiments.

Limitations

  • Stated: the origin of xenosiderophore substrates used by non-siderophore producers such as Candida and Cryptococcus remains unresolved, and further studies are required to elucidate Sit1 siderophore recognition, uptake mechanism, and intracellular fate.
  • Stated: mouse and human macrophage immunobiology differs, including controversy over functional iNOS in human macrophages, which motivated testing both J774A.1 and U937 cells.
  • Evident from the text: the study relies on macrophage cell lines and ex vivo primary mouse macrophages, not an in vivo mammalian infection model, and phagosomal iron status is inferred from SIT1 expression, ferritin levels, and survival phenotypes rather than directly measured.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1371/journal.ppat.1001322

Cites

53 of 53 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 35 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    Higher hemin concentrations were required for slower growth of C. glabrata due to hemin toxicity when compared to C. albicans.

    “higher hemin concentrations were required for a slower growth of C. glabrata due to hemin toxicity when compared to C. albicans”
    Quote in the paperNot a substantial claim
  • Findingown data

    Computational topological analyses of Sit1 identified sequence signatures characteristic of members of Major Facilitator Superfamily of transporters.

    “Computational topological analyses of Sit1 identified sequence signatures characteristic of members of the Major Facilitator Superfamily of transporters”
    Quote in the paperNot a substantial claim
  • Findingown data

    Sit1 functions as a determinant for C. glabrata to survive macrophage killing in a manner dependent on macrophage iron status.

    “Sit1 functions as a determinant for C. glabrata to survive macrophage killing in a manner that is dependent on macrophage iron status.”
    Quote in the paperSubstantial claim
  • Findingown data

    Using macrophage models of human iron overload disease, C. glabrata senses altered iron levels within the phagosomal compartment.

    “Using macrophage models of human iron overload disease, we demonstrate that C. glabrata senses altered iron levels within the phagosomal compartment.”
    Quote in the paperSubstantial claim
  • Findingown data

    Protein BLAST analysis of a 51 amino acid sequence comprising last extra-cellular loop (residues 531 to 581) exclusively returned known siderophore transporters and uncharacterized proteins predicted to encode fungal siderophore transporters.

    “Protein BLAST analysis [25] of a 51 amino acid sequence comprising the last extra-cellular loop (from amino acid residue 531 to 581) exclusively returned known siderophore transporters and uncharacterized proteins predicted to encode fungal siderophore transporters”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [25] SF Altschul, 1997, Gapped BLAST and PSI-BLAST: a new generation of protein database search programs., Nucleic Acids Res source

      Not checked in the free run.

  • Classified as probably not a substantial claim. 27
  • May combine two claims. 31

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data

    This SIderophore Transporter Domain (SITD) is conserved in many pathogenic fungi.

    “This SIderophore Transporter Domain (SITD) is conserved in many pathogenic fungi”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    C. albicans was able to efficiently utilize heme as source of Fe at sub-micromolar hemin concentrations, whereas C. glabrata required 100-fold higher hemin concentrations to achieve same level of growth.

    “whereas C. albicans was able to efficiently utilize heme as a source of Fe at sub-micromolar hemin concentrations, C. glabrata required 100-fold higher hemin concentrations to achieve the same level of growth.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    No significant differences between wild type, sit1Δ and sit1Δ::SIT1 strains were observed in CFU 2 hours post-infection in presence or absence of ferrichrome, strongly suggesting equally efficient phagocytosis.

    “CFU were monitored 2 hours post-infection and no significant differences between the wild type, sit1Δ and sit1Δ::SIT1 strains were observed in the presence or absence of ferrichrome, strongly suggesting equally efficient phagocytosis of the three yeast strains by macrophages”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data

    Sit1 orthologues predicted across ascomycetes, and to lesser extent basidiomycetes, share high sequence identity within carboxyl terminus of protein.

    “Sit1 orthologues predicted to exist across the ascomycetes, and to lesser extent basidiomycetes, share high sequence identity within the carboxyl terminus of the protein.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Sit1 protein strongly accumulates at plasma membrane under Fe deficiency in absence of siderophore substrate and this accumulation is largely unaltered by addition of ferrichrome.

    “indirect immunofluorescence as well as live microscopy of C. glabata Sit1 revealed strong accumulation of the Sit1 protein at the plasma membrane under Fe deficiency in the absence of siderophore substrate and this accumulation was largely unaltered by the addition of ferrichrome”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Steady state SIT1 mRNA levels are elevated under Fe deficiency and this elevation is sustained throughout time-course over 6 hours.

    “Figure 2B shows that steady state SIT1 mRNA levels are elevated under Fe deficiency and that this elevation is sustained throughout the time-course of the experiment over 6 hours.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingcitation

    C. glabrata genome encodes a single open reading frame, CAGL0E04092g, encoding a predicted membrane protein with 14 transmembrane domains and high degree of identity (72%) to siderophore-Fe transporter Arn1, designated SIT1.

    “Analysis of the Candida glabrata genome [24] predicts the existence of orthologues to the Saccharomyces cerevisiae high affinity reductive Fe assimilation pathway as well as a single open reading frame, CAGL0E04092g, encoding a predicted membrane protein with 14 transmembrane domains and a high degree of identity (72%) to the siderophore-Fe transporter, Arn1, that we have designated SIT1”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [24] DJ Sherman, 2009, Genolevures: protein families and synteny among complete hemiascomycetous yeast proteomes and genomes., Nucleic Acids Res source

      Not checked in the free run.

  • May combine two claims.

    Findingreasoning

    Incubation with hepcidin increased ferritin levels in cells expressing wild type FPN, as predicted from Fpn internalization and degradation, resulting in increased cellular Fe accumulation.

    “Incubation with hepcidin increased ferritin levels in cells expressing wild type FPN, as would be predicted from Fpn internalization and degradation, resulting in increased cellular Fe accumulation”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Pre-loading U937 cells with Fe by culturing in presence of 20 μM FAC promoted growth of C. glabrata and abrogated requirement for Sit1-mediated siderophore utilization in enhanced C. glabrata survival.

    “Pre-loading U937 cells with Fe by culturing in the presence of 20 µM FAC promoted the growth of C. glabrata and abrogated the requirement for Sit1-mediated siderophore utlization in enhanced C. glabrata survival”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    A strain carrying the SIT1Y575A allele integrated at the endogenous genomic locus was strongly impaired in its ability to use ferrichrome as an Fe source despite exhibiting levels of protein expression and plasma membrane localization comparable to the wild type strain.

    “A strain carrying the SIT1Y575A allele integrated at the endogenous genomic locus (Figure S5C) was strongly impaired in its ability to use ferrichrome as an Fe source ( Figure 7A ) despite exhibiting levels of protein expression (Figure S5D) and plasma membrane localization ( Figure 7B and Figure S5E) comparable to the wild type strain.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Plasma membrane accumulation of Flag antibody was observed in non-permeabilized cells expressing carboxyl-loop-Flag Sit1, and this signal was absent in cells expressing vector alone.

    “As shown in Figure S5B, plasma membrane accumulation of the Flag antibody was observed in non-permeabilized cells expressing carboxyl-loop-Flag Sit1. This immunofluorescence signal was absent in cells expressing vector alone”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Low macrophage Fe levels associated with FpnC326Y expression impose Fe deficiency on phagocytosed C. glabrata cells, as demonstrated by elevated SIT1 expression and increased dependence on ferrichrome and Sit1 for proliferation.

    “the low macrophage Fe levels associated with FpnC326Y expression impose Fe deficiency on phagocytosed C. glabrata cells, as demonstrated by elevated SIT1 expression and further supported by the increased dependence on ferrichrome and Sit1 for proliferation.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Neither the sit1Δ nor the strain bearing the Sit1Y575A substitution in Sit1 showed the Sit1-mediated siderophore-dependent increase in the ability to survive macrophage killing.

    “In contrast to wild type C. glabrata cells, neither the sit1Δ nor the strain bearing the Sit1Y575A substitution in Sit1 showed the Sit1-mediated siderophore-dependent increase in the ability to survive macrophage killing”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Exposure to hepcidin leads to predominantly intracellular localization of wild type protein, whereas hepcidin-insensitive FpnC326Y protein remains at cell surface.

    “Exposure to hepcidin leads to a predominantly intracellular localization of the wild type protein, whereas the hepcidin-insensitive FpnC326Y protein remains at the cell surface.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    SIT1 expression is low under Fe repletion and shows robust mRNA and protein accumulation under Fe deficiency.

    “SIT1 characterization demonstrated that expression is responsive to environmental Fe bioavailability, showing low levels of expression under Fe repletion and robust mRNA and protein accumulation under Fe deficiency ( Figure 2B and 2C ).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Both wild type and FpnC326Y-expressing cells, but not cells expressing FpnH32R, show plasma membrane ferroportin accumulation in absence of hepcidin.

    “Both the wild type and FpnC326Y-expressing cells, but not cells expressing FpnH32R, show plasma membrane ferroportin accumulation in the absence of hepcidin”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    These siderophores are either not transported by Sit1 or C. glabrata lacks a mechanism of releasing Fe from these siderophores intracellularly.

    “suggesting that these siderophores are either not transported by Sit1 or C. glabrata lacks a mechanism of releasing the Fe from these siderophores intracellularly.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    Mammalian cellular immunity relies heavily on action of macrophages, which also play critical role in systemic Fe homeostasis.

    “Mammalian cellular immunity relies heavily on the action of macrophages, cells that also play a critical role in systemic Fe homeostasis [13].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [13] NC Andrews, 2008, Forging a field: the golden age of iron biology., Blood source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Given weak hemolytic activity and inability to efficiently import heme, C. glabrata relies primarily on circulating host Fe sources to satisfy its requirements for Fe.

    “Thus, given its weak hemolytic activity and inability to efficiently import heme, we predict that C. glabrata relies primarily on circulating host Fe sources to satisfy its requirements for Fe.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    Fe2+ can generate damaging reactive oxygen species via Fenton/Haber Weiss chemistry.

    “Fe2+ also has the potential to generate damaging reactive oxygen species (ROS) via Fenton/Haber Weiss chemistry [5]”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [5] B Halliwell, 1985, The importance of free radicals and catalytic metal ions in human diseases., Mol Aspects Med source

      Not checked in the free run.

  • May combine two claims.

    Premisenone given

    Macrophages mediate microbial containment through localized reduction of serum transferrin-bound Fe, decreased Fe export and depletion of Fe from macrophage phagosome.

    “A key mechanism through which macrophages mediate microbial containment is through the localized reduction of serum transferrin-bound Fe, decreased Fe export and the depletion of Fe from the macrophage phagosome”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Free Fe3+ concentration is below 10−9 M at pH 7 and free serum Fe concentration is approximately 10−24 M.

    “Given the low Fe bioavailability in an aqueous environment (free Fe3+ concentration below 10−9 M at pH 7) and the further strict active withholding of Fe within mammalian hosts (free serum Fe concentration of approximately 10−24 M) [9]”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [9] KN Raymond, 2003, Enterobactin: an archetype for microbial iron transport., Proc Natl Acad Sci U S A source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    C. glabrata has emerged as an opportunistic fungal pathogen that causes life-threatening infectious disease in humans.

    “Candida glabrata has emerged as an opportunistic fungal pathogen that causes life-threatening infectious disease in humans [1].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [1] C Moran, 2010, Comparison of costs, length of stay, and mortality associated with <italic>Candida glabrata</italic> and <italic>Candida albicans</italic> bloodstream infections., Am J Infect Control source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    These observations are consistent with reports of greater microbial burden in patients with Fe overload disease and predict that mutations affecting macrophage Fe levels may compromise microbial containment and dictate dependence of microbes on distinct mechanisms of Fe uptake.

    “These observations are consistent with reports of greater microbial burden in patients with Fe overload disease [15], [18], [19], [20], [21] and predict that mutations affecting macrophage Fe levels may not only compromise microbial containment, but may also dictate the dependence of microbes on distinct mechanisms of Fe uptake.”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [15] FA Khan, 2007, Association of hemochromatosis with infectious diseases: expanding spectrum., Int J Infect Dis source

      Not checked in the free run.

    • cites [18] JC Barton, 2009, Hemochromatosis and <italic>Vibrio vulnificus</italic> wound infections., J Clin Gastroenterol source

      Not checked in the free run.

    • cites [19] S Cunningham-Rundles, 2000, Effect of transfusional iron overload on immune response., J Infect Dis source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    CaSit1 is required for invasion and penetration of human epithelial cells in an in vitro model of oral candidiasis but was not required for murine systemic candida dissemination.

    “CaSit1 has been shown to be required for the invasion and penetration of human epithelial cells in an in vitro model of oral candidiasis but was not required for murine systemic candida dissemination [11]”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [11] P Heymann, 2002, The siderophore iron transporter of <italic>Candida albicans</italic> (Sit1p/Arn1p) mediates uptake of ferrichrome-type siderophores and is required for epithelial invasion., Infect Immun source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    S. cerevisiae possesses two high-affinity mechanisms of Fe acquisition.

    “Eukaryotic Fe homeostasis has best been described in the budding yeast, Saccharomyces cerevisiae that possesses two high-affinity mechanisms of Fe acquisition [7].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [7] CC Philpott, 2008, Response to iron deprivation in <italic>Saccharomyces cerevisiae</italic>., Eukaryot Cell source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    This is consistent with high levels of labile intracellular Fe in FPNH32R macrophages that elevate abundance of ferritin and might be inefficiently withheld from C. glabrata.

    “This is consistent with the high levels of labile intracellular Fe in FPNH32R macrophages that elevate the abundance of ferritin and might be inefficiently withheld from C. glabrata.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Reductive Fe uptake from ferric citrate was downregulated in the presence of serum, as evidenced by decreased Fe uptake and reduced ferrireductase activity, whereas that from ferrichrome-type siderophores was elevated.

    “The results obtained showed that reductive Fe uptake from ferric citrate was downregulated in the presence of serum, as evidenced by decreased Fe uptake and reduced ferrireductase activity, whereas that from ferrichrome-type siderophores was elevated.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [43] E Lesuisse, 2002, Siderophore uptake by <italic>Candida albicans</italic>: effect of serum treatment and comparison with <italic>Saccharomyces cerevisiae</italic>., Yeast source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Individuals with malignant hematological disorders, such as acute myeloid leukemia, sustain increased mycoses, especially by Candida and Aspergilli.

    “Individuals with malignant hematological disorders, such as acute myeloid leukemia, who typically present with very high serum Fe, serum ferritin and transferrin saturation, are known to sustain increased mycoses, especially by Candida and Aspergilli [38], [39].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [38] C Iglesias-Osma, 1995, Iron metabolism and fungal infections in patients with haematological malignancies., J Clin Pathol source

      Not checked in the free run.

    • cites [39] JE Karp, 1986, Association of reduced total iron binding capacity and fungal infections in leukemic granulocytopenic patients., J Clin Oncol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Fe overload correlates with poor prognosis in patients infected with HIV.

    “and correlates with poor prognosis and in patients infected with HIV, a virus known to depend on and manipulate target cellular Fe accumulation [40].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [40] H Drakesmith, 2008, Viral infection and iron metabolism., Nat Rev Microbiol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Transfusion-dependent thalassemia patients present with increased susceptibility to widespread microbial infection that correlates with the degree of Fe overload.

    “In the same way, transfusion-dependent thalassemia patients present with increased susceptibility to widespread microbial infection that correlates with the degree of Fe overload [19].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [19] S Cunningham-Rundles, 2000, Effect of transfusional iron overload on immune response., J Infect Dis source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    Our results underscore a role for siderophore utilization in the successful existence of C. glabrata as a human commensal as well as during pathogenesis, primarily with respect to survival to phagocytosis by mammalian macrophages.

    “Our results underscore a role for siderophore utilization in the successful existence of C. glabrata as a human commensal as well as during pathogenesis, primarily with respect to survival to phagocytosis by mammalian macrophages.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Siderophore utilization is critical to microbial colonization of healthy immunocompetent individuals in its state of commensalism.

  • May combine two claims.

    Interpretationown data

    Macrophage Fe-withholding from phagocytosed microorganisms is fundamental for containment of microbial growth and impacts mechanisms available for microbial iron uptake, imposing strict dependence of C. glabrata on availability and utilization of xenosiderophores via Sit1.

    “Moreover, these results indicate that macrophage Fe-withholding from phagocytosed microorganisms is fundamental for the containment of microbial growth and also impacts on the mechanisms available for microbial iron uptake, imposing a strict dependence of C. glabrata on the availability and utilization of xenosiderophores via Sit1.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    The flatiron (ffe) mouse model of human Ferroportin Disease harbors the FPNH32R mutation.

    “the flatiron (ffe) mouse model of human Ferroportin Disease, which harbors the FPNH32R mutation [36].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [36] IE Zohn, 2007, The flatiron mutation in mouse ferroportin acts as a dominant negative to cause ferroportin disease., Blood source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    The decreased dependence of C. glabrata on siderophore-Fe, decreased SIT1 expression indicative of increased Fe satiety, increased C. glabrata fitness and increased levels of host cell ferritin expression result from the inability of Fe-laden macrophages to withhold essential metal from internalized C. glabrata cells.

    “We infer that this results from the inability of Fe-laden macrophages to withhold this essential metal from internalized C. glabrata cells given the decreased dependence of C. glabrata on siderophore-Fe, decreased SIT1 expression indicative of increased Fe satiety, increased C. glabrata fitness and the increased levels of host cell ferritin expression.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    This is consistent with refractility of FpnC326Y to hepcidin-stimulated degradation and Fe levels in these macrophages that are lower than wild type.

    “This is consistent with the refractility of FpnC326Y to hepcidin-stimulated degradation and Fe levels in these macrophages that are lower than wild type.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation

    Modest increase in ferritin levels may reflect contribution of hepcidin-sensitive endogenous wild type homodimers that correctly localize to plasma membrane.

    “Given that Fpn functions as a dimer [37], this modest increase in ferritin levels may reflect the contribution of hepcidin-sensitive endogenous wild type homodimers that correctly localize to the plasma membrane.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [37] I De Domenico, 2007, Evidence for the multimeric structure of ferroportin., Blood source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Clinical correlations exist between elevated Fe bioavailability and increased microbial burden, extending beyond diseases of Fe overload, including higher susceptibility to Vibrio vulnificus infection.

    “The existence of clinical correlations between elevated Fe bioavailability and increased microbial burden extend beyond diseases of Fe overload for which the higher susceptibility to bacterial infection, notably to Vibrio vulnificus, has been described [15], [18].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [15] FA Khan, 2007, Association of hemochromatosis with infectious diseases: expanding spectrum., Int J Infect Dis source

      Not checked in the free run.

    • cites [18] JC Barton, 2009, Hemochromatosis and <italic>Vibrio vulnificus</italic> wound infections., J Clin Gastroenterol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Fe overload is an independent risk factor for post-transplant infection.

    “clinical evidence also indicates that Fe overload is an independent risk factor for post-transplant infection [20], [21]”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [20] N Singh, 2008, Iron overload and unique susceptibility of liver transplant recipients to disseminated disease due to opportunistic pathogens., Liver Transpl source

      Not checked in the free run.

    • cites [21] DP Kontoyiannis, 2007, Increased bone marrow iron stores is an independent risk factor for invasive aspergillosis in patients with high-risk hematologic malignancies and recipients of allogeneic hematopoietic stem cell transplantation., Cancer source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The FPNC326Y disease-associated mutation results in a similar clinical outcome to that observed in patients carrying HFE mutations present in approximately 80% of HH patients and exhibiting an allelic frequency of over 10% in Caucasians of European descent.

    “The FPNC326Y disease-associated mutation results in a similar clinical outcome to that observed in patients carrying HFE mutations present in approximately 80% of HH patients and exhibiting an allelic frequency of over 10% in Caucasians of European descent [13].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [13] NC Andrews, 2008, Forging a field: the golden age of iron biology., Blood source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Each experiment included 3 replicates per experimental sample.

    “Each experiment included 3 replicates per experimental sample.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    Localized siderophore reservoirs are likely available to C. glabrata and other non-siderophore producers.

    “Given the ubiquitous nature of siderophore production and the mixed nature of the mammalian microbiome localized siderophore reservoirs are likely available to C. glabrata and other non-siderophore producers.”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: Siderophore production is ubiquitous in nature.
    • Rests on an assumption: The mammalian microbiome is mixed.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Conidia of a siderophore-deficient mutant exhibit decreased levels of stored Fe and higher sensitivity to oxidative stress.

    “the conidia of a siderophore-deficient mutant exhibit decreased levels of stored Fe and higher sensitivity to oxidative stress [47].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [47] M Schrettl, 2007, Distinct roles for intra- and extracellular siderophores during <italic>Aspergillus fumigatus</italic> infection., PLoS Pathog source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Aspergillus fumigatus synthesizes chemically distinct siderophores for the solubilization of extracellular Fe as well as for its safe intracellular storage.

    “Aspergillus fumigatus, one of the most prevalent air-borne human pathogens, synthesizes chemically distinct siderophores for the solubilization of extracellular Fe as well as for its safe intracellular storage [47].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [47] M Schrettl, 2007, Distinct roles for intra- and extracellular siderophores during <italic>Aspergillus fumigatus</italic> infection., PLoS Pathog source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    SIT1 expression levels were normalized against those of ACT1.

    “SIT1 expression levels were normalized against those of ACT1.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The routing of trans-Golgi Arn1 away from the plasma membrane and to the vacuolar protein-sorting pathway requires the monomeric clathrin adaptor, Gga2.

    “The routing of trans-Golgi Arn1 away from the plasma membrane and to the vacuolar protein-sorting pathway requires the monomeric clathrin adaptor, Gga2 [50].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [50] Y Deng, 2009, Gga2 mediates sequential ubiquitin-independent and ubiquitin-dependent steps in the trafficking of ARN1 from the trans-Golgi network to the vacuole., J Biol Chem source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Each experiment included 6 replicates per experimental sample.

    “Each experiment included 6 replicates per experimental sample.”
    • Also stated in: § Integrity of the Sit1 SITD is critical for ferrichrome utilization · plain
      “Each experiment included 6 replicates per experimental sample.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    A recent study identified a complex and dynamic profile of fungal species in the oral cavity of healthy individuals, the number of which parallels that observed for bacteria.

    “A recent study by Ghannoum et al. characterizing the mycobiome of the oral cavity of healthy individuals identified in this host milieu a complex and dynamic profile of fungal species, the number of which parallels that observed for bacteria [46].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [46] MA Ghannoum, Characterization of the oral fungal microbiome (mycobiome) in healthy individuals., PLoS Pathog source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    C. albicans Sit1 as well as S. cerevisiae Enb1 localize predominantly to the plasma membrane in either the absence or presence of substrate siderophore.

    “Interestingly, C. albicans Sit1 as well as S. cerevisiae Enb1 [7], also localize predominantly to the plasma membrane in either the absence or presence of substrate siderophore [51].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [7] CC Philpott, 2008, Response to iron deprivation in <italic>Saccharomyces cerevisiae</italic>., Eukaryot Cell source

      Not checked in the free run.

    • cites [51] CJ Hu, 2002, Characterization and functional analysis of the siderophore-iron transporter CaArn1p in <italic>Candida albicans</italic>., J Biol Chem source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductown data

    Results are representative of at least 3 experiments.

    “Results are shown as mean survival compared to the wild type grown in the absence of ferrichrome supplementation and are representative of at least 3 experiments.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Siderophore utilization by Aspergillus fumigatus is required for virulence, in contrast to reductive Fe assimilation.

    “siderophore utilization by this saprophytic fungus has been shown to be required for virulence, in contrast to reductive Fe assimilation [48]”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [48] M Schrettl, 2004, Siderophore biosynthesis but not reductive iron assimilation is essential for <italic>Aspergillus fumigatus</italic> virulence., J Exp Med source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    An alpha-Pgk1 antibody was used as loading control.

    “An alpha-Pgk1 antibody was used as loading control.”
    Quote in the paperNot a substantial claim
52 of 52

  • The paper relies on this without support.

    The mouse J774A.1 and human U937 macrophage cell lines are valid models of human macrophage iron homeostasis and microbicidal activity.

    “Using the mouse macrophage-like cell line, J774A.1, isogenic wild type, sit1Δ and sit1Δ::SIT1 C. glabrata strains were grown under Fe limiting conditions”
    “The human monocytic cell line, U937, was differentiated into mature macrophages prior to infection with the indicated C. glabrata strains grown under Fe deficiency without or with supplementation of 10 µM FC and yeast survival evaluated after 18, 24 and 36 hours of infection.”

  • The paper relies on this without support.

    Siderophore concentrations in serum are negligible.

    “possibly reflecting the sterile environment of the serum in which siderophore concentrations are predicted to be negligible.”

  • The paper relies on this without support.

    C. glabrata cannot efficiently use heme.

    “Given the inability of C. glabrata to efficiently use heme”

  • The paper relies on this without support.

    Siderophore production is ubiquitous in nature.

    “Given the ubiquitous nature of siderophore production”

  • The paper relies on this without support.

    The mammalian microbiome is mixed.

    “and the mixed nature of the mammalian microbiome”

  • The paper relies on this without support.

    The severe Fe restriction imposed by mammalian hosts as an antimicrobial strategy dictates that Fe acquisition is a bona fide virulence determinant.

    “The severe Fe restriction imposed by mammalian hosts as an antimicrobial strategy dictates that Fe acquisition is a bona fide virulence determinant.”

  • The paper relies on this without support.

    J774A.1 and U937 are well-established macrophage cell lines.

    “using well-established macrophage cell lines”

  • The paper relies on this without support.

    Siderophore utilization is critical to microbial colonization of healthy immunocompetent individuals in its state of commensalism.

    “Whilst siderophore utilization is also undoubtedly critical to microbial colonization of healthy immunocompetent individuals in its state of commensalism”

  • The paper relies on this without support.

    Sit1 shares high sequence identity with S. cerevisiae Arn1 that translates into similar siderophore substrate specificity, including ferrichromes and coprogen.

    “Sit1 is syntenic to its orthologous gene in S. cerevisiae, Arn1, with which it shares high sequence identity that translates into similar siderophore substrate specificity, which includes ferrichromes and coprogen.”
9 of 9

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • The sole xenosiderophore transporter CgSit1 typifies the siderophore-mediated iron uptake system in C. glabrata.

    “The sole xenosiderophore transporter CgSit1 typifies the siderophore-mediated iron uptake system in C. glabrata [123].”

    From Candida glabrata: A Lot More Than Meets the Eye (2019) · cites this paper as [123]

  • CgSit1 is pivotal to survival in the iron-limited host environment.

    “CgSit1 has been shown to be pivotal to survival in the iron-limited host environment [123].”

    From Candida glabrata: A Lot More Than Meets the Eye (2019) · cites this paper as [123]

  • C. glabrata is not known to use hemoglobin or transferrin for iron acquisition, in contrast to C. albicans.

    “Especially for C. glabrata, which (in contrast to C. albicans) is not known to use hemoglobin or transferrin for iron acquisition (8, 15)”

    From A novel hybrid iron regulation network combines features from pathogenic and nonpathogenic yeasts (2016) · cites this paper as [15]

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Related

Authors
Nevitt, T., & Thiele, D. J.
Affiliations
Duke Medical Center (US)
Corresponding author
Thiele, Dennis J.
ORCID
Thiele, Dennis J.
Venue
PLoS Pathogens
Publisher
Public Library of Science (PLoS)
Published
2011-03-17
Volume
7
Issue
3
Pages
e1001322
ISSN
  • 1553-7366 (ISSN-L)
  • 1553-7374
DOI
10.1371/journal.ppat.1001322
PMC
PMC3060170
OpenAlex
W2035619455
Semantic Scholar
ad15bf7a5ebedecef3ec7eb92404d3ea40cb5a9c
Cited by
95
References
53
Citations by year
2026: 5 · 2025: 4 · 2024: 6 · 2023: 4 · 2022: 3 · 2021: 6 · 2020: 12 · 2019: 7 · 2018: 3 · 2017: 7 · 2016: 9 · 2015: 5 · 2014: 6 · 2013: 11 · 2012: 4
FWCI
5.18
Type
journal article
Language
en
Open access
Licence
Locations
  • PLoS Pathogens (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
  • Figshare (submittedVersion, cc-by)
Funders
  • Flatiron Health
  • National Institutes of Health
  • National Institute of General Medical Sciences (GM 41840, R01GM041840)
Topics
  • Health Sciences › Medicine › Infectious Diseases › Antifungal resistance and susceptibility
  • Health Sciences › Medicine › Epidemiology › Fungal Infections and Studies
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
Keywords
Candida glabrata, siderophore-mediated iron acquisition, macrophage killing, iron overload, fungal virulence, host-pathogen interaction, iron deficiency
MeSH terms
Amino Acid Sequence, Candidiasis, Carrier Proteins, Fungal Proteins, Humans, Iron, Molecular Sequence Data, Siderophores, Virulence Factors, Candida glabrata

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-09.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-09.

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