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The evolution of drug resistance in clinical isolates of Candida albicans.

Ford CB, Funt JM, Abbey D, Issi L, Guiducci C, Martinez DA, Delorey T, Li BY, White TC, Cuomo C, Rao RP, Berman J, Thompson DA, Regev A - Elife (2015)

Bottom Line: Studies in clinical isolates have implicated multiple mechanisms in resistance, but have focused on large-scale aberrations or candidate genes, and do not comprehensively chart the genetic basis of adaptation.LOH events were commonly associated with acquired resistance, and SNPs in 240 genes may be related to host adaptation.Conversely, most aneuploidies were transient and did not correlate with drug resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Broad Institute of MIT and Harvard, Cambridge, United States.

ABSTRACT
Candida albicans is both a member of the healthy human microbiome and a major pathogen in immunocompromised individuals. Infections are typically treated with azole inhibitors of ergosterol biosynthesis often leading to drug resistance. Studies in clinical isolates have implicated multiple mechanisms in resistance, but have focused on large-scale aberrations or candidate genes, and do not comprehensively chart the genetic basis of adaptation. Here, we leveraged next-generation sequencing to analyze 43 isolates from 11 oral candidiasis patients. We detected newly selected mutations, including single-nucleotide polymorphisms (SNPs), copy-number variations and loss-of-heterozygosity (LOH) events. LOH events were commonly associated with acquired resistance, and SNPs in 240 genes may be related to host adaptation. Conversely, most aneuploidies were transient and did not correlate with drug resistance. Our analysis also shows that isolates also varied in adherence, filamentation, and virulence. Our work reveals new molecular mechanisms underlying the evolution of drug resistance and host adaptation.

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Analysis of discordant sites.(A) Degree of concordance (Y axis) with Sequenom iPlexgenotyping for 1973 SNP X strain combinations overall (leftmost red bar;93.9%) and in each tested strain (X axis). (B) Shown are theclasses of discordant sites by genotype as defined by Illumina (orange)or Sequenome (teal) (X axis) and the prevalence (Y axis) of that genotypecall in Sequenom (blue) and Illumina (orange) based discordant calls. Themost common discrepancies arose when Sequenom typing classified a site ashomozygous, but Illumina sequencing identified it as heterozygous.(C–G) Comparison on distributions ofquality features between concordant (blue bars) and discordant (greenbars) sites: (C) depth of coverage, (D) RMSMapping Quality (MQ) score, (E) PHRED scaled quality scorefor each base call, shown as log-normalized ‘QUAL’ scores,(F) quality by depth (QD) score for each variant site,and (G) the allele balance ratio (AB Score) for each variantsite.DOI:http://dx.doi.org/10.7554/eLife.00662.005
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fig1s1: Analysis of discordant sites.(A) Degree of concordance (Y axis) with Sequenom iPlexgenotyping for 1973 SNP X strain combinations overall (leftmost red bar;93.9%) and in each tested strain (X axis). (B) Shown are theclasses of discordant sites by genotype as defined by Illumina (orange)or Sequenome (teal) (X axis) and the prevalence (Y axis) of that genotypecall in Sequenom (blue) and Illumina (orange) based discordant calls. Themost common discrepancies arose when Sequenom typing classified a site ashomozygous, but Illumina sequencing identified it as heterozygous.(C–G) Comparison on distributions ofquality features between concordant (blue bars) and discordant (greenbars) sites: (C) depth of coverage, (D) RMSMapping Quality (MQ) score, (E) PHRED scaled quality scorefor each base call, shown as log-normalized ‘QUAL’ scores,(F) quality by depth (QD) score for each variant site,and (G) the allele balance ratio (AB Score) for each variantsite.DOI:http://dx.doi.org/10.7554/eLife.00662.005

Mentions: We sequenced the genomic DNA of the isolates as well as the C.albicans lab strain, SC5314, using Illumina sequencing (53-283X coverage,103X on average, ‘Materials and methods’, Table 1) and identified in each series point mutations, LOHevents and aneuploidies that were not present in the first strain in that series. Byconvention, all mutations were defined relative to SC5314, the C.albicans genome reference strain. We validated our pipeline for detectionof point mutations using Sequenom iPlex genotyping (Storm et al., 2003) (‘Materials and methods’). Weinterrogated 1973 SNPs in 27 isolates from nine clinical series and found that theiPlex base calls matched 1853 (93.9%, Figure1—figure supplement 1A, Table2) of the calls from our computational analysis of the sequencing data.Evaluation of the discordant sites showed somewhat lower quality scores by certainmetrics but did not identify any metrics that could be used to systematically revisefiltering in our computational pipeline without a radical reduction in sensitivity(Figure 1—figure supplement1B–G).10.7554/eLife.00662.009Table 2.


The evolution of drug resistance in clinical isolates of Candida albicans.

Ford CB, Funt JM, Abbey D, Issi L, Guiducci C, Martinez DA, Delorey T, Li BY, White TC, Cuomo C, Rao RP, Berman J, Thompson DA, Regev A - Elife (2015)

Analysis of discordant sites.(A) Degree of concordance (Y axis) with Sequenom iPlexgenotyping for 1973 SNP X strain combinations overall (leftmost red bar;93.9%) and in each tested strain (X axis). (B) Shown are theclasses of discordant sites by genotype as defined by Illumina (orange)or Sequenome (teal) (X axis) and the prevalence (Y axis) of that genotypecall in Sequenom (blue) and Illumina (orange) based discordant calls. Themost common discrepancies arose when Sequenom typing classified a site ashomozygous, but Illumina sequencing identified it as heterozygous.(C–G) Comparison on distributions ofquality features between concordant (blue bars) and discordant (greenbars) sites: (C) depth of coverage, (D) RMSMapping Quality (MQ) score, (E) PHRED scaled quality scorefor each base call, shown as log-normalized ‘QUAL’ scores,(F) quality by depth (QD) score for each variant site,and (G) the allele balance ratio (AB Score) for each variantsite.DOI:http://dx.doi.org/10.7554/eLife.00662.005
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Related In: Results  -  Collection

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fig1s1: Analysis of discordant sites.(A) Degree of concordance (Y axis) with Sequenom iPlexgenotyping for 1973 SNP X strain combinations overall (leftmost red bar;93.9%) and in each tested strain (X axis). (B) Shown are theclasses of discordant sites by genotype as defined by Illumina (orange)or Sequenome (teal) (X axis) and the prevalence (Y axis) of that genotypecall in Sequenom (blue) and Illumina (orange) based discordant calls. Themost common discrepancies arose when Sequenom typing classified a site ashomozygous, but Illumina sequencing identified it as heterozygous.(C–G) Comparison on distributions ofquality features between concordant (blue bars) and discordant (greenbars) sites: (C) depth of coverage, (D) RMSMapping Quality (MQ) score, (E) PHRED scaled quality scorefor each base call, shown as log-normalized ‘QUAL’ scores,(F) quality by depth (QD) score for each variant site,and (G) the allele balance ratio (AB Score) for each variantsite.DOI:http://dx.doi.org/10.7554/eLife.00662.005
Mentions: We sequenced the genomic DNA of the isolates as well as the C.albicans lab strain, SC5314, using Illumina sequencing (53-283X coverage,103X on average, ‘Materials and methods’, Table 1) and identified in each series point mutations, LOHevents and aneuploidies that were not present in the first strain in that series. Byconvention, all mutations were defined relative to SC5314, the C.albicans genome reference strain. We validated our pipeline for detectionof point mutations using Sequenom iPlex genotyping (Storm et al., 2003) (‘Materials and methods’). Weinterrogated 1973 SNPs in 27 isolates from nine clinical series and found that theiPlex base calls matched 1853 (93.9%, Figure1—figure supplement 1A, Table2) of the calls from our computational analysis of the sequencing data.Evaluation of the discordant sites showed somewhat lower quality scores by certainmetrics but did not identify any metrics that could be used to systematically revisefiltering in our computational pipeline without a radical reduction in sensitivity(Figure 1—figure supplement1B–G).10.7554/eLife.00662.009Table 2.

Bottom Line: Studies in clinical isolates have implicated multiple mechanisms in resistance, but have focused on large-scale aberrations or candidate genes, and do not comprehensively chart the genetic basis of adaptation.LOH events were commonly associated with acquired resistance, and SNPs in 240 genes may be related to host adaptation.Conversely, most aneuploidies were transient and did not correlate with drug resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Broad Institute of MIT and Harvard, Cambridge, United States.

ABSTRACT
Candida albicans is both a member of the healthy human microbiome and a major pathogen in immunocompromised individuals. Infections are typically treated with azole inhibitors of ergosterol biosynthesis often leading to drug resistance. Studies in clinical isolates have implicated multiple mechanisms in resistance, but have focused on large-scale aberrations or candidate genes, and do not comprehensively chart the genetic basis of adaptation. Here, we leveraged next-generation sequencing to analyze 43 isolates from 11 oral candidiasis patients. We detected newly selected mutations, including single-nucleotide polymorphisms (SNPs), copy-number variations and loss-of-heterozygosity (LOH) events. LOH events were commonly associated with acquired resistance, and SNPs in 240 genes may be related to host adaptation. Conversely, most aneuploidies were transient and did not correlate with drug resistance. Our analysis also shows that isolates also varied in adherence, filamentation, and virulence. Our work reveals new molecular mechanisms underlying the evolution of drug resistance and host adaptation.

Show MeSH
Related in: MedlinePlus