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Cited by 35Antifungal resistance and susceptibilityEvolution and Genetic DynamicsFungal and yeast genetics research

Acquisition of cross-azole tolerance and aneuploidy in Candida albicans strains evolved to posaconazole

Open access
Kukurudz, Rebekah J, Chapel, Madison, Wonitowy, Quinn, Adamu Bukari, Abdul-Rahman, Sidney, Brooke, Sierhuis, Riley, Gerstein, Aleeza C·2022·G3
DOI 10.1093/g3journal/jkac156PMC PMC9434289

Published version of a preprint

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Abstract

A number of in vitro studies have examined the acquisition of drug resistance to the triazole fluconazole, a first-line treatment for many Candida infections. Much less is known about posaconazole, a newer triazole. We conducted the first in vitro experimental evolution of replicates from 8 diverse strains of Candida albicans in a high level of the fungistatic drug posaconazole. Approximately half of the 132 evolved replicates survived 50 generations of evolution, biased toward some of the strain backgrounds. We found that although increases in drug resistance were rare, increases in drug tolerance (the slow growth of a subpopulation of cells in a level of drug above the resistance level) were common across strains. We also found that adaptation to posaconazole resulted in widespread cross-tolerance to other azole drugs. Widespread aneuploidy was observed in evolved replicates from some strain backgrounds. Trisomy of at least one of chromosomes 3, 6, and R was identified in 11 of 12 whole-genome sequenced evolved SC5314 replicates. These findings document rampant evolved cross-tolerance among triazoles and highlight that increases in drug tolerance can evolve independently of drug resistance in a diversity of C. albicans strain backgrounds.

AI snapshot

Written from the paper’s full text.

TL;DR

This study performed the first in vitro experimental evolution of 8 diverse *Candida albicans* strains in high posaconazole, finding that survival was common only with 72 h transfers and that increased drug tolerance and cross-tolerance to other azoles evolved frequently while increased resistance was rare. Evolved SC5314 replicates often acquired aneuploidies, especially ChrR trisomy, but links between specific aneuploidies and azole phenotypes remained unclear.

Key findings

  • Of 132 POS0.5-evolved replicates, none survived the 24 h transfer experiment, while 67 of 132 survived the 72 h transfer experiment; survival varied by strain background and was not significantly correlated with ancestral growth ability in drug (Pearson’s correlation, t6 = 0.91, P = 0.40).
  • Increases in drug resistance were rare; instead, replicates from 6 strain backgrounds evolved increased susceptibility, while tolerance increased in 6 strain backgrounds. Median tolerance increases were GC75: 34% POS / 26% FLC; SC5314: 37% POS / 35% FLC; P78048: 24% POS / 43% FLC.
  • Adaptation to posaconazole produced widespread cross-tolerance to other azoles: nearly all evolved SC5314 replicates showed increased tolerance to all 5 azole drugs tested (POS, FLC, VCZ, CTR, MCZ) and essentially no change in NYT or 5-FC.
  • Aneuploidy was widespread: 11 of 12 whole-genome sequenced evolved SC5314 replicates had trisomy of at least one of chromosomes 3, 6, and R; ChrR trisomy was most common, in 9 replicates. No clear clustering linked specific aneuploidies to azole resistance or tolerance phenotypes.

Methods

  • Eight clinical *C. albicans* strains from different phylogenetic clades were used; 12 single-colony replicates per strain were evolved in YPD + 0.5 µg/mL posaconazole (POS0.5) or YPD alone, with 1:1,000 serial transfers every 24 or 72 h for 5 transfers (~50 generations). In total, 132 replicates were evolved per transfer duration in POS and 96 per transfer duration in YPD.
  • Drug susceptibility and tolerance were measured by disk diffusion assays and quantified with diskImageR as RAD20 (resistance) and FoG20 (tolerance) for POS and FLC across all replicates, plus additional drugs for SC5314; growth ability was measured as OD600 at 24 and 72 h.
  • Flow cytometry assessed genome size/ploidy, and whole-genome sequencing was performed on 12 evolved SC5314 replicates plus 2 ancestral replicates, followed by variant calling and YMAP analysis for copy number variation and loss of heterozygosity.

Limitations

  • The paper states that it cannot differentiate whether aneuploidies provide increased tolerance to POS0.5 (i.e. are beneficial) or are a consequence of azole exposure (i.e. are neutral), and no clear phenotypic pattern separated strains with ChrR or other trisomies; additional work is required to link aneuploidy to phenotype.
  • The paper notes that some trisomies had copy numbers between 2 and 3, likely indicating polymorphic populations, and it cannot rule out that some aneuploidies were unstable and lost during growth from frozen culture; small CNVs had no obvious adaptive benefit and were likely mapping artifacts.
  • Evident from the text: whole-genome karyotype analysis was limited to SC5314, the 24 h transfer experiment had no survivors, and several strain backgrounds had too few surviving replicates for statistical testing.

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

Integrity signals

Registry

Crossref · 10.1093/g3journal/jkac156

Cites

86 of 86 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 40 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    Replicate survival to 72 h transfers in POS0.5 was not equal among strain backgrounds.

    “Replicate survival to 72 h transfers in POS0.5 was not equal among strain backgrounds”
    Quote in the paperSubstantial claim
  • Findingown data

    Evolved replicates from GC75 had no improvement over ancestral replicates at 24 or 72 h in POS.

    “Evolved replicates from GC75 had no improvement over ancestral replicates at 24 or 72 h.”
    Quote in the paperNot a substantial claim
  • Findingown data

    Six replicates from 4 strains (P75016, GC75, SC5314, and P78048) acquired decreased susceptibility (increased resistance) to POS.

    “Six replicates from 4 strains did deviate from the rest and acquired decreased susceptibility (increased resistance; P75016, GC75, SC5314, and P78048).”
    Quote in the paperSubstantial claim
  • Findingown data

    No replicates survived in the POS0.5 24 h transfer experiment.

    “No replicates survived in the POS0.5 24 h transfer experiment”
    Quote in the paperSubstantial claim
  • Findingown data

    In the 72 h transfer POS0.5 evolution experiment, 67 of 132 evolved replicates survived to the end.

    “approximately half of the replicates (67 of 132) survived to the end of the POS0.5 72 h transfer experiment”
    • Also stated in: § Abstract · hedged
      “Approximately half of the 132 evolved replicates survived 50 generations of evolution, biased toward some of the strain backgrounds.”
    Quote in the paperNot a substantial claim
  • May combine two claims. 28
  • The quote only partly states this claim. 2
  • Classified as probably not a substantial claim. 19

  • May combine two claims.

    Findingown data

    Evolved replicates from P78048, SC5314, and FH1 had a minor reduction in growth ability in YPD at 24 h, which continued to 72 h for SC5314 and FH1.

    “Evolved replicates from 3 strains (P78048, SC5314, and FH1) had a minor reduction in growth ability in YPD at 24 h, which continued to 72 h for SC5314 and FH1.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Although a small number of replicates evolved increased resistance to FLC, the majority of replicates in most backgrounds increased in susceptibility to FLC.

    “although a small number of replicates evolved an increased resistance to FLC, the majority of replicates in most backgrounds increased in susceptibility”
    Quote in the paperSubstantial claim

  • The quote only partly states this claim.

    Findingown data

    There was no correlation between growth improvements in POS and growth reductions in YPD in any strain background at either 24 h or 72 h.

    “There was no correlation between growth improvements in POS and growth reductions in YPD in any strain background at either 24 h”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Evolved replicates from 6 strain backgrounds increased in tolerance to POS, while 2 of the strains with a higher number of surviving replicates did not change.

    “Evolved replicates from 6 strain backgrounds increased in tolerance, while 2 of the strains with a higher number of surviving replicates did not change”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The median increase in tolerance among replicates in POS and FLC was 34% and 26% for GC75, 37% and 35% for SC5314, and 24% and 43% for P78048.

    “The median increase in tolerance among replicates was quite high and fairly similar in both POS and FLC for GC75 (POS: 34%, FLC: 26%), SC5314 (POS: 37%, FLC: 35%), and P78048 isolates (POS: 24%, FLC: 43%).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Evolved replicates tended to increase in FLC susceptibility (decrease in resistance) yet increase in FLC tolerance.

    “evolved replicates tended to increase in FLC susceptibility (decrease in resistance) yet increase in FLC tolerance.”
    Quote in the paperSubstantial claim

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

    Findingown data

    T101 had a minor improvement in growth ability at 24 h in YPD.

    “T101 had a minor improvement in growth ability at 24 h in YPD.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Evolved replicates from strain GC75 in the 24 h YPD transfer experiment decreased in tolerance, while replicates from strain FH1 in the 24 h YPD transfer experiment increased in tolerance.

    “Evolved replicates from strain GC75 in the 24 h transfer experiment decreased in tolerance, while replicates from strain FH1 in the 24 h transfer experiment increased in tolerance.”
    Quote in the paperNot a substantial claim

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

    Findingnone given

    All replicate lines survived 24 and 72 h transfers in the standard rich medium YPD.

    “All replicate lines survived 24 and 72 h transfers in the standard rich medium YPD.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Two additional evolved SC5314 replicates had an extra copy of Chr6, one alone and one in tandem with an extra copy of Chr3.

    “Two additional evolved replicates also had an extra copy of Chr6, one alone and one in tandem with an extra copy of Chr3”
    Quote in the paperNot a substantial claim

  • May combine two claims. The quote only partly states this claim.

    Findingown data

    Replicates evolved with 24 and 72 h transfers in YPD exhibited much smaller changes in resistance and tolerance than POS-evolved replicates, though some evolved strains did differ significantly compared to the ancestors.

    “Replicates that were evolved with 24 and 72 h transfers in YPD exhibited much smaller changes in resistance and tolerance, though some evolved strains did differ significantly compared to the ancestors”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Of the 9 replicates with ChrR trisomy, one had additional aneuploidies in Chr3 and Chr6, and a second had an extra copy of Chr4.

    “Of these 9, one replicate had additional aneuploidies in Chr3 and Chr6, and a second replicate had an extra copy of Chr4.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Nearly all evolved SC5314 replicates showed increased tolerance to all 5 azole drugs examined and essentially no change in NYT or 5-FC.

    “nearly all evolved replicates showed increased tolerance to all 5 azole drugs examined and essentially no change in NYT or 5-FC.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In YPD with 24 h transfers, replicates from one strain background significantly increased in tolerance while replicates from a second significantly decreased.

    “We found a similar result here in YPD when replicates were transferred for 24 h: replicates from one strain background significantly increased in tolerance while replicates from a second significantly decreased.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The top 2 principal components of the 10 azole phenotypes had eigenvalues above 1 and combined to explain 77% of the variance.

    “The top 2 principal components both had eigenvalues above 1 and combined to explain 77% of the variance.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    These aneuploidies were concurrent with increases in tolerance to POS and other azoles, but not increases in resistance to POS or other tested drug classes.

    “These aneuploidies were concurrent with increases in tolerance to POS (and other azoles), but not increases in resistance to POS or other tested drug classes.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The majority of surviving replicates increased in POS tolerance and acquired cross-tolerance to FLC and other azole drugs.

    “The majority of surviving replicates did increase in POS tolerance and acquired cross-tolerance to FLC and other azole drugs.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    PC2 (13.1% of the variance, eigenvalue = 1.1) differentiated among the different azole drugs, with VCZ most similar to POS.

    “PC2 (13.1% of the variance, eigenvalue = 1.1) differentiated among the different azole drugs, demonstrating that VCZ was most similar to POS”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The only other mutations in the targeted genes were 7 LOH events in S4 upstream and downstream of GZF3, interspersed with heterozygous positions, so not indicative of a large-scale LOH event.

    “The only other mutations in these genes were 7 LOH events in positions in S4 upstream and downstream of GZF3 (at base pairs 646493, 646559, 646560, 646608, 647030, 649877, and 649963) that were interspersed with heterozygous positions (so not indicative of a large-scale LOH event).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Strain backgrounds with fewer surviving replicates were more likely to increase in POS drug tolerance and more likely to increase in genome size.

    “Strain backgrounds with fewer surviving replicates were more likely to increase in POS drug tolerance and more likely to increase in genome size.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The majority of evolved replicates improved in rapid growth in the evolutionary level of drug, yet very few replicates increased in POS resistance.

    “The majority of evolved replicates improved in rapid growth in the evolutionary level of drug, yet very few replicates increased in POS resistance.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data

    The ancestral and evolved replicates of T101 are resistant to FLC, and changes to FLC tolerance in evolved replicates cannot be examined in this framework.

    “The ancestral (and evolved) replicates of T101 are resistant to FLC and changes to FLC tolerance in evolved replicates cannot be examined in this framework.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    Tolerance is either more evolvable than resistance or was the primary phenotype under selection during the in vitro evolution to POS0.5.

    “suggesting that tolerance is either more evolvable than resistance or was the primary phenotype under selection during the in vitro evolution to POS0.5.”
    Quote in the paperSubstantial claim

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

    Premisecitation

    Candida albicans is the primary species responsible for mucosal disease.

    “Candida albicans the primary species responsible for mucosal disease”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Bongomin et al. 2017 source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    All ancestral strains were initially diploid.

    “all were initially diploid”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    A small number of localized CNVs were present in evolved lines, but none have an obvious adaptive benefit.

    “A small number of localized CNV were also present in evolved lines, but none have an obvious adaptive benefit”
    Quote in the paperNot a substantial claim

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

    Premisecitation

    Fungal tolerance may play a role in persistent candidemia and mortality.

    “it may play a role in persistent candidemia (Rosenberg et al. 2018) and mortality (Levinson et al. 2021).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Rosenberg et al. 2018 source

      Not checked in the free run.

    • cites Levinson et al. 2021 source

      Not checked in the free run.

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

    Premisecitation

    A recent screen of 235 clinical Candida spp. isolates found that resistant isolates also tended to be more tolerant to the fungistatic drug fluconazole.

    “A recent screen of 235 clinical Candida spp. isolates found that resistant isolates also tended to be more tolerant to the fungistatic drug fluconazole (FLC; Salama and Gerstein 2022).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Salama and Gerstein 2022

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Premisecitation

    Multiple ERG11 mutations are required for POS resistance.

    “it seems that multiple ERG11 mutations are required for POS resistance (Li et al. 2004).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Li et al. 2004

      Not checked in the free run.

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

    Premisecitation

    Extra copies of Chr3, Chr4, and Chr5 have previously been shown to confer increased resistance to FLC.

    “Extra copies of hr3 (Perepnikhatka et al. 1999; Ford et al. 2015), Chr4 (Perepnikhatka et al. 1999; Anderson et al. 2017), and Chr5 (Coste et al. 2006; Selmecki et al. 2006, 2008; Ford et al. 2015; Todd et al. 2019) have previously been shown to confer increased resistance to FLC”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Perepnikhatka et al. 1999 source

      Not checked in the free run.

    • cites Ford et al. 2015 source

      Not checked in the free run.

    • cites Anderson et al. 2017 source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    The genetic basis of major adaptive pathways in FLC-resistant C. albicans involves alterations or overexpression of ERG11 and upregulation of drug efflux pumps encoded by CDR1, CDR2, TAC1, MRR1, and MDR1.

    “the genetic basis of major adaptive pathways have been identified: the first involves alterations or overexpression of ERG11, which produces the target demethylase”
    • Also stated in: § Introduction · plain
      “the second is through the upregulation of drug efflux pumps encoded by CDR1, CDR2, TAC1, MRR1, and MDR1”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Asai et al. 1999 source

      Not checked in the free run.

    • cites Lamb et al. 2000 source

      Not checked in the free run.

    • cites Selmecki et al. 2008 source

      Not checked in the free run.

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

    Premisecitation

    A large-scale in vitro evolution experiment in FLC found that changes in tolerance evolved independently of resistance.

    “a large-scale in vitro evolution experiment in FLC found that changes in tolerance evolved independently of resistance (Gerstein and Berman 2020).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Gerstein and Berman 2020

      Not checked in the free run.

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

    Premisecitation

    The azole drugs FLC and voriconazole are first-line treatments for candidiasis and aspergillosis, respectively.

    “The azole drugs FLC and voriconazole (VCZ) are first-line treatments for candidiasis and aspergillosis, respectively (Pappas et al. 2016; Patterson et al. 2016).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Pappas et al. 2016 source

      Not checked in the free run.

    • cites Patterson et al. 2016 source

      Not checked in the free run.

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

    Premisecitation

    Posaconazole effectively treated infections resistant to first-line azoles in A. fumigatus, C. albicans, and C. neoformans.

    “One of the newest azoles, posaconazole (POS), effectively treated infections resistant to first-line azoles in A. fumigatus, C. albicans, and C. neoformans (Chau et al. 2004; Xiao et al. 2004; Firinu et al. 2011; Sionov et al. 2012).”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Chau et al. 2004 source

      Not checked in the free run.

    • cites Xiao et al. 2004 source

      Not checked in the free run.

    • cites Firinu et al. 2011 source

      Not checked in the free run.

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

    Premisecitation

    Chr6 and ChrR were previously found to be the 2 most common aneuploidies remaining in otherwise euploid strains after 28 days of daily passaging of initially tetraploid and initially aneuploid strains in standard lab YPD.

    “Interestingly, Chr6 and ChrR were previously found to be the 2 most common aneuploidies remaining in otherwise euploid strains after 28 days of daily passaging of initially tetraploid and initially aneuploid strains in standard lab YPD (Hickman et al. 2015).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Hickman et al. 2015 source

      Not checked in the free run.

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

    Premisecitation

    Chr3 aneuploidy was previously identified to confer an increase in FLC tolerance, due at least in part to an extra copy of the urea transporter NPR2.

    “Chr3 aneuploidy was previously identified to confer an increase in FLC tolerance, due at least in part to an extra copy of the urea transporter NPR2 (Mount et al. 2018).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Mount et al. 2018 source

      Not checked in the free run.

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

    Premisecitation

    Laboratory evolution of microbes at high population sizes is a particularly powerful way to examine parallelism and constraint in evolution at genomic and phenotypic levels.

    “Laboratory evolution of microbes at high population sizes (at times termed “experimental evolution” or “adaptive laboratory evolution”) is a particularly powerful way to examine parallelism and constraint in evolution at genomic and phenotypic levels (Cooper 2018; Gerstein and Sharp 2021).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Cooper 2018 source

      Not checked in the free run.

    • cites Gerstein and Sharp 2021

      Not checked in the free run.

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

    Premisecitation

    Transcriptomics and phenotypic analyses demonstrated that all Rim proteins, including RIM9 on ChrR, were important for FLC tolerance.

    “Transcriptomics and phenotypic analyses demonstrated that all Rim proteins (including RIM9, on ChrR) were important for FLC tolerance (Garnaud et al. 2018).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Garnaud et al. 2018 source

      Not checked in the free run.

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

    Premisecitation

    GZF3, a GATA-type transcription factor of unknown function, was one of 2 genes, alongside CRZ1, identified in an overexpression screen of 572 genes for FLC tolerance.

    “Also intriguing and worthy of further study is GZF3, a GATA-type transcription factor of unknown function, one of 2 genes (alongside CRZ1) identified in an overexpression screen of 572 genes for FLC tolerance (Delarze et al. 2020).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Delarze et al. 2020 source

      Not checked in the free run.

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

    Premisecitation

    A previous study in a different strain background found that an extra copy of ChrR increased resistance to FLC, KCZ, and MCZ.

    “a previous study in a different strain background found that an extra copy of ChrR increased resistance to FLC, KCZ, and MCZ (Li et al. 2015).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Li et al. 2015

      Not checked in the free run.

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

    Premisecitation

    Changes in chromosome copy number are frequently observed in experimental studies of fungal microbes, yet they are often observed without identifying a directly mechanistic link to fitness.

    “Changes in chromosome copy number are frequently observed in experimental studies of fungal microbes, yet they are often observed without identifying a directly mechanistic link to fitness (Gerstein and Sharp 2021).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Gerstein and Sharp 2021

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    In vitro evolution to antifungal drugs may not always result in increased drug resistance, and different suites of mutations may confer a benefit to different levels of the same drug.

    “Combined, these results suggest that in vitro evolution to antifungal drugs may not always result in increased drug resistance and that different suites of mutations may confer a benefit to different levels of the same drug.”
    Quote in the paperSubstantial claim

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

    Premisecitation

    Experimental laboratory evolution has been an effective method to study pathways of adaptation in a variety of biological contexts.

    “Experimental laboratory evolution has been an effective method to study pathways of adaptation in a variety of biological contexts (Kawecki et al. 2012; Cooper 2018).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Kawecki et al. 2012 source

      Not checked in the free run.

    • cites Cooper 2018 source

      Not checked in the free run.

  • May combine two claims.

    Interpretationcitation

    There is potential for drug tolerance to evolve quickly, which may have important clinical implications, particularly in persistent candidemia infections when populations are exposed to drug stress for long periods while the underlying strain remains drug susceptible.

    “It seems clear that there is potential for drug tolerance to evolve quickly, which may have important clinical implications, particularly in cases of persistent candidemia infections when populations are exposed to drug stress for long periods of time, yet the underlying strain remains drug susceptible (Rosenberg et al. 2018).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Rosenberg et al. 2018 source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    The aneuploidies observed in POS-evolved lines are largely different from those shown to confer increased FLC resistance, indicating that the genetic pathway to acquiring POS tolerance is distinct.

    “these are largely different from those that have been shown to confer an increase in resistance to the related azole drug FLC, indicating that the genetic pathway to acquiring POS tolerance is distinct.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Tolerance has both a genetic component, likely involving genes in membrane biosynthesis/integrity and stress response pathways, and an environmental component, as growth conditions have been linked to the degree of tolerance exhibited.

    “Tolerance has both a genetic component, likely involving genes in membrane biosynthesis/integrity and the stress response pathways (Cowen et al. 2014; Rosenberg et al. 2018; Berman and Krysan 2020; Todd and Selmecki 2020) and an environmental component, as growth conditions have been linked to the degree of tolerance exhibited (Gerstein et al. 2016; Berman and Krysan 2020).”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Cowen et al. 2014 source

      Not checked in the free run.

    • cites Rosenberg et al. 2018 source

      Not checked in the free run.

    • cites Berman and Krysan 2020

      Not checked in the free run.

46 of 46

  • The paper relies on this without support.

    Posaconazole is a fungistatic drug.

    “in a high level of the fungistatic drug posaconazole”
    “in the fungistatic drug POS”

  • The paper relies on this without support.

    Whether consistent POS exposure also selects for beneficial aneuploidy has not been determined.

    “Whether consistent POS exposure also selects for beneficial aneuploidy has not been determined.”

  • The paper relies on this without support.

    Very few studies have directly examined how different conditions influence how readily tolerance evolves in C. albicans.

    “Very few studies have (yet) directly examined how different conditions influence how readily tolerance evolves in C. albicans.”

  • The paper relies on this without support.

    The rapid evolution of tolerance and cross-tolerance through in vitro evolution has not been previously observed in azoles nor linked to aneuploidy.

    “The rapid evolution of tolerance and cross-tolerance through in vitro evolution has not been previously observed in azoles nor linked to aneuploidy.”

  • The paper relies on this without support.

    The 14 genes analyzed were previously implicated in antifungal resistance.

    “We conducted a targeted genomic analysis of 14 genes previously implicated in antifungal resistance: TAC1, GSL2, CDR1, CDR2, ERG11, CRZ1, MRR1, UPC2, MDR1, RRP6, RIM9, NPR2, and GZF3.”
5 of 5

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?

  • Azole tolerance and cross-tolerance to different azole drugs can evolve rapidly in vitro.

    “Intriguingly, our recent experiments demonstrate that azole tolerance and cross-tolerance to different azole drugs can evolve rapidly in vitro (Todd and Selmecki 2020; Kukurudz et al. 2022), yet the mechanisms that distinguish drug-resistant and drug-tolerant phenotypes in fungi are not fully characterized.”

    From Antifungal Drug Concentration Impacts the Spectrum of Adaptive Mutations in <i>Candida albicans</i> (2023) · cites this paper as Kukurudz et al. 2022

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Posaconazole disk dose is overstated 1,000-fold

    Minor

    Four microliters of a 0.625 mg/mL solution contains 0.0025 mg (2.5 μg), not 2.5 mg.

    Reported 2.5 · recomputed 0.0025

  • Targeted gene list contains 13 genes, not 14

    Minor

    The stated total is 14, but the accompanying list names only 13 genes.

  • PCA eigenvalues and variance percentages disagree

    Minor

    The two principal components must use the same total variance. PC1's figures imply a total eigenvalue of about 3.91, whereas PC2's imply about 8.40.

3 of 3

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Authors
Kukurudz, R. J., Chapel, M., Wonitowy, Q., Adamu Bukari, A.-R., Sidney, B., Sierhuis, R., & Gerstein, A. C.
Affiliations
  • Department of Microbiology, The University of Manitoba , Winnipeg, MB R3T 2N2, Canada
  • University of Manitoba (CA)
Corresponding author
Gerstein, Aleeza C
ORCID
Venue
G3
Publisher
Oxford University Press (OUP)
Published
2022-07-26
Volume
12
Issue
9
ISSN
2160-1836 (ISSN-L)
DOI
10.1093/g3journal/jkac156
PMC
PMC9434289
OpenAlex
W4288038380
Semantic Scholar
500b25485ab83175870aa4b8b3617c623a297388
Cited by
35
References
86
Citations by year
2026: 3 · 2025: 9 · 2024: 12 · 2023: 7 · 2022: 4
FWCI
3.90
Type
journal article
Language
en
Open access
Licence
Locations
  • G3 Genes Genomes Genetics (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
  • NSERC Discovery Grant
  • University of Manitoba and a University of Manitoba University Research Grants Program
  • University of Manitoba, Faculty of Science
  • NSERC Undergraduate Student Research Awards
  • EvoFunPath
Topics
  • Health Sciences › Medicine › Infectious Diseases › Antifungal resistance and susceptibility
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Evolution and Genetic Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
Keywords
Candida albicans, posaconazole, drug resistance, drug tolerance, aneuploidy, experimental evolution, trisomy, SC5314
MeSH terms
Aneuploidy, Antifungal Agents, Azoles, Candida albicans, Microbial Sensitivity Tests, Triazoles, Fluconazole, Drug Resistance, Fungal

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.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

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