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One small step for a yeast--microevolution within macrophages renders Candida glabrata hypervirulent due to a single point mutation.

Brunke S, Seider K, Fischer D, Jacobsen ID, Kasper L, Jablonowski N, Wartenberg A, Bader O, Enache-Angoulvant A, Schaller M, d'Enfert C, Hube B - PLoS Pathog. (2014)

Bottom Line: Continuous co-incubation of C. glabrata with a murine macrophage cell line for over six months resulted in a striking alteration in fungal morphology: The growth form changed from typical spherical yeasts to pseudohyphae-like structures - a phenotype which was stable over several generations without any selective pressure.Similarly, the Evo mutant significantly increased TNFα production in the brain on day 2, which is mirrored in macrophages confronted with the Evo mutant, but not with the parental wild type.These results indicate that microevolutionary processes in host-simulative conditions can elicit adaptations of C. glabrata to distinct host niches and even lead to hypervirulent strains.

View Article: PubMed Central - PubMed

Affiliation: Integrated Research and Treatment Center, Sepsis und Sepsisfolgen, Center for Sepsis Control and Care (CSCC), Universitätsklinikum Jena, Jena, Germany; Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology - Hans Knoell Institute Jena (HKI), Jena, Germany.

ABSTRACT
Candida glabrata is one of the most common causes of candidemia, a life-threatening, systemic fungal infection, and is surpassed in frequency only by Candida albicans. Major factors contributing to the success of this opportunistic pathogen include its ability to readily acquire resistance to antifungals and to colonize and adapt to many different niches in the human body. Here we addressed the flexibility and adaptability of C. glabrata during interaction with macrophages with a serial passage approach. Continuous co-incubation of C. glabrata with a murine macrophage cell line for over six months resulted in a striking alteration in fungal morphology: The growth form changed from typical spherical yeasts to pseudohyphae-like structures - a phenotype which was stable over several generations without any selective pressure. Transmission electron microscopy and FACS analyses showed that the filamentous-like morphology was accompanied by changes in cell wall architecture. This altered growth form permitted faster escape from macrophages and increased damage of macrophages. In addition, the evolved strain (Evo) showed transiently increased virulence in a systemic mouse infection model, which correlated with increased organ-specific fungal burden and inflammatory response (TNFα and IL-6) in the brain. Similarly, the Evo mutant significantly increased TNFα production in the brain on day 2, which is mirrored in macrophages confronted with the Evo mutant, but not with the parental wild type. Whole genome sequencing of the Evo strain, genetic analyses, targeted gene disruption and a reverse microevolution experiment revealed a single nucleotide exchange in the chitin synthase-encoding CHS2 gene as the sole basis for this phenotypic alteration. A targeted CHS2 mutant with the same SNP showed similar phenotypes as the Evo strain under all experimental conditions tested. These results indicate that microevolutionary processes in host-simulative conditions can elicit adaptations of C. glabrata to distinct host niches and even lead to hypervirulent strains.

No MeSH data available.


Related in: MedlinePlus

Differences in the host cytokine response to wild type and Evo strains in vivo and in vitro.A. Selected cytokines and MPO levels were measured in murine organs at day 2 after infection with wild type or Evo strain. Significantly higher cytokine levels were found specifically in the brain of mice infected with the Evo strain, reflecting the transient high fungal burden in this organ. B. Release of the same cytokines by murine RAW246.7 macrophages was tested for the different strains. Only strains bearing the point mutation in the CHS2 gene (Evo and CHSEvo) elicited an increased release of TNFα.
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ppat-1004478-g005: Differences in the host cytokine response to wild type and Evo strains in vivo and in vitro.A. Selected cytokines and MPO levels were measured in murine organs at day 2 after infection with wild type or Evo strain. Significantly higher cytokine levels were found specifically in the brain of mice infected with the Evo strain, reflecting the transient high fungal burden in this organ. B. Release of the same cytokines by murine RAW246.7 macrophages was tested for the different strains. Only strains bearing the point mutation in the CHS2 gene (Evo and CHSEvo) elicited an increased release of TNFα.

Mentions: To explain the differences in murine virulence, we measured cytokine and myeloperoxidase levels as markers for inflammation in the different organs. In all organs with similar fungal burden of wild type and evolved strain, we found the tissue concentrations of these markers to be indistinguishable. Yet strikingly, the evolved strain induced significantly increased levels of the proinflammatory cytokines TNFα and IL-6 specifically in the brain at day two, mirroring the high fungal burden in this organ (Fig. 5A). Similarly, IL-1β and the inflammation marker MPO were found at increased levels, albeit not statistically significant. This increased inflammation also coincided with the most severe clinical symptoms. At later time points, the brain cytokine levels of mice infected with either strain became similar again, paralleling the fungal burden of the evolved and parental strain in this organ. We concluded that the transiently increased virulence of the evolved strain was likely caused at least in part by a strongly increased inflammatory response in the brain.


One small step for a yeast--microevolution within macrophages renders Candida glabrata hypervirulent due to a single point mutation.

Brunke S, Seider K, Fischer D, Jacobsen ID, Kasper L, Jablonowski N, Wartenberg A, Bader O, Enache-Angoulvant A, Schaller M, d'Enfert C, Hube B - PLoS Pathog. (2014)

Differences in the host cytokine response to wild type and Evo strains in vivo and in vitro.A. Selected cytokines and MPO levels were measured in murine organs at day 2 after infection with wild type or Evo strain. Significantly higher cytokine levels were found specifically in the brain of mice infected with the Evo strain, reflecting the transient high fungal burden in this organ. B. Release of the same cytokines by murine RAW246.7 macrophages was tested for the different strains. Only strains bearing the point mutation in the CHS2 gene (Evo and CHSEvo) elicited an increased release of TNFα.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4214790&req=5

ppat-1004478-g005: Differences in the host cytokine response to wild type and Evo strains in vivo and in vitro.A. Selected cytokines and MPO levels were measured in murine organs at day 2 after infection with wild type or Evo strain. Significantly higher cytokine levels were found specifically in the brain of mice infected with the Evo strain, reflecting the transient high fungal burden in this organ. B. Release of the same cytokines by murine RAW246.7 macrophages was tested for the different strains. Only strains bearing the point mutation in the CHS2 gene (Evo and CHSEvo) elicited an increased release of TNFα.
Mentions: To explain the differences in murine virulence, we measured cytokine and myeloperoxidase levels as markers for inflammation in the different organs. In all organs with similar fungal burden of wild type and evolved strain, we found the tissue concentrations of these markers to be indistinguishable. Yet strikingly, the evolved strain induced significantly increased levels of the proinflammatory cytokines TNFα and IL-6 specifically in the brain at day two, mirroring the high fungal burden in this organ (Fig. 5A). Similarly, IL-1β and the inflammation marker MPO were found at increased levels, albeit not statistically significant. This increased inflammation also coincided with the most severe clinical symptoms. At later time points, the brain cytokine levels of mice infected with either strain became similar again, paralleling the fungal burden of the evolved and parental strain in this organ. We concluded that the transiently increased virulence of the evolved strain was likely caused at least in part by a strongly increased inflammatory response in the brain.

Bottom Line: Continuous co-incubation of C. glabrata with a murine macrophage cell line for over six months resulted in a striking alteration in fungal morphology: The growth form changed from typical spherical yeasts to pseudohyphae-like structures - a phenotype which was stable over several generations without any selective pressure.Similarly, the Evo mutant significantly increased TNFα production in the brain on day 2, which is mirrored in macrophages confronted with the Evo mutant, but not with the parental wild type.These results indicate that microevolutionary processes in host-simulative conditions can elicit adaptations of C. glabrata to distinct host niches and even lead to hypervirulent strains.

View Article: PubMed Central - PubMed

Affiliation: Integrated Research and Treatment Center, Sepsis und Sepsisfolgen, Center for Sepsis Control and Care (CSCC), Universitätsklinikum Jena, Jena, Germany; Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology - Hans Knoell Institute Jena (HKI), Jena, Germany.

ABSTRACT
Candida glabrata is one of the most common causes of candidemia, a life-threatening, systemic fungal infection, and is surpassed in frequency only by Candida albicans. Major factors contributing to the success of this opportunistic pathogen include its ability to readily acquire resistance to antifungals and to colonize and adapt to many different niches in the human body. Here we addressed the flexibility and adaptability of C. glabrata during interaction with macrophages with a serial passage approach. Continuous co-incubation of C. glabrata with a murine macrophage cell line for over six months resulted in a striking alteration in fungal morphology: The growth form changed from typical spherical yeasts to pseudohyphae-like structures - a phenotype which was stable over several generations without any selective pressure. Transmission electron microscopy and FACS analyses showed that the filamentous-like morphology was accompanied by changes in cell wall architecture. This altered growth form permitted faster escape from macrophages and increased damage of macrophages. In addition, the evolved strain (Evo) showed transiently increased virulence in a systemic mouse infection model, which correlated with increased organ-specific fungal burden and inflammatory response (TNFα and IL-6) in the brain. Similarly, the Evo mutant significantly increased TNFα production in the brain on day 2, which is mirrored in macrophages confronted with the Evo mutant, but not with the parental wild type. Whole genome sequencing of the Evo strain, genetic analyses, targeted gene disruption and a reverse microevolution experiment revealed a single nucleotide exchange in the chitin synthase-encoding CHS2 gene as the sole basis for this phenotypic alteration. A targeted CHS2 mutant with the same SNP showed similar phenotypes as the Evo strain under all experimental conditions tested. These results indicate that microevolutionary processes in host-simulative conditions can elicit adaptations of C. glabrata to distinct host niches and even lead to hypervirulent strains.

No MeSH data available.


Related in: MedlinePlus