RESUMO
Evolutionary changes in the hepatitis B virus (HBV) genome could reflect its adaptation to host-induced selective pressure. Leveraging paired human exome and ultra-deep HBV genome-sequencing data from 567 affected individuals with chronic hepatitis B, we comprehensively searched for the signatures of this evolutionary process by conducting "genome-to-genome" association tests between all human genetic variants and viral mutations. We identified significant associations between an East Asian-specific missense variant in the gene encoding the HBV entry receptor NTCP (rs2296651, NTCP S267F) and mutations within the receptor-binding region of HBV preS1. Through in silico modeling and in vitro preS1-NTCP binding assays, we observed that the associated HBV mutations are in proximity to the NTCP variant when bound and together partially increase binding affinity to NTCP S267F. Furthermore, we identified significant associations between HLA-A variation and viral mutations in HLA-A-restricted T cell epitopes. We used in silico binding prediction tools to evaluate the impact of the associated HBV mutations on HLA presentation and observed that mutations that result in weaker binding affinities to their cognate HLA alleles were enriched. Overall, our results suggest the emergence of HBV escape mutations that might alter the interaction between HBV PreS1 and its cellular receptor NTCP during viral entry into hepatocytes and confirm the role of HLA class I restriction in inducing HBV epitope variations.
Assuntos
Vírus da Hepatite B , Mutação , Transportadores de Ânions Orgânicos Dependentes de Sódio , Simportadores , Humanos , Vírus da Hepatite B/genética , Transportadores de Ânions Orgânicos Dependentes de Sódio/genética , Transportadores de Ânions Orgânicos Dependentes de Sódio/metabolismo , Simportadores/genética , Simportadores/metabolismo , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Hepatite B Crônica/virologia , Hepatite B Crônica/genética , Genoma Viral , Antígenos de Superfície da Hepatite B/genética , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Genômica/métodos , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/metabolismoRESUMO
Population isolates such as those in Finland benefit genetic research because deleterious alleles are often concentrated on a small number of low-frequency variants (0.1% ≤ minor allele frequency < 5%). These variants survived the founding bottleneck rather than being distributed over a large number of ultrarare variants. Although this effect is well established in Mendelian genetics, its value in common disease genetics is less explored1,2. FinnGen aims to study the genome and national health register data of 500,000 Finnish individuals. Given the relatively high median age of participants (63 years) and the substantial fraction of hospital-based recruitment, FinnGen is enriched for disease end points. Here we analyse data from 224,737 participants from FinnGen and study 15 diseases that have previously been investigated in large genome-wide association studies (GWASs). We also include meta-analyses of biobank data from Estonia and the United Kingdom. We identified 30 new associations, primarily low-frequency variants, enriched in the Finnish population. A GWAS of 1,932 diseases also identified 2,733 genome-wide significant associations (893 phenome-wide significant (PWS), P < 2.6 × 10-11) at 2,496 (771 PWS) independent loci with 807 (247 PWS) end points. Among these, fine-mapping implicated 148 (73 PWS) coding variants associated with 83 (42 PWS) end points. Moreover, 91 (47 PWS) had an allele frequency of <5% in non-Finnish European individuals, of which 62 (32 PWS) were enriched by more than twofold in Finland. These findings demonstrate the power of bottlenecked populations to find entry points into the biology of common diseases through low-frequency, high impact variants.
Assuntos
Doença , Frequência do Gene , Fenótipo , Humanos , Pessoa de Meia-Idade , Doença/genética , Estônia , Finlândia , Frequência do Gene/genética , Predisposição Genética para Doença/genética , Estudo de Associação Genômica Ampla , Metanálise como Assunto , Reino Unido , População Branca/genéticaRESUMO
Common single-nucleotide polymorphisms (SNPs) are predicted to collectively explain 40-50% of phenotypic variation in human height, but identifying the specific variants and associated regions requires huge sample sizes1. Here, using data from a genome-wide association study of 5.4 million individuals of diverse ancestries, we show that 12,111 independent SNPs that are significantly associated with height account for nearly all of the common SNP-based heritability. These SNPs are clustered within 7,209 non-overlapping genomic segments with a mean size of around 90 kb, covering about 21% of the genome. The density of independent associations varies across the genome and the regions of increased density are enriched for biologically relevant genes. In out-of-sample estimation and prediction, the 12,111 SNPs (or all SNPs in the HapMap 3 panel2) account for 40% (45%) of phenotypic variance in populations of European ancestry but only around 10-20% (14-24%) in populations of other ancestries. Effect sizes, associated regions and gene prioritization are similar across ancestries, indicating that reduced prediction accuracy is likely to be explained by linkage disequilibrium and differences in allele frequency within associated regions. Finally, we show that the relevant biological pathways are detectable with smaller sample sizes than are needed to implicate causal genes and variants. Overall, this study provides a comprehensive map of specific genomic regions that contain the vast majority of common height-associated variants. Although this map is saturated for populations of European ancestry, further research is needed to achieve equivalent saturation in other ancestries.
Assuntos
Estatura , Mapeamento Cromossômico , Polimorfismo de Nucleotídeo Único , Humanos , Estatura/genética , Frequência do Gene/genética , Genoma Humano/genética , Estudo de Associação Genômica Ampla , Haplótipos/genética , Desequilíbrio de Ligação/genética , Polimorfismo de Nucleotídeo Único/genética , Europa (Continente)/etnologia , Tamanho da Amostra , FenótipoRESUMO
We assembled an ancestrally diverse collection of genome-wide association studies (GWAS) of type 2 diabetes (T2D) in 180,834 affected individuals and 1,159,055 controls (48.9% non-European descent) through the Diabetes Meta-Analysis of Trans-Ethnic association studies (DIAMANTE) Consortium. Multi-ancestry GWAS meta-analysis identified 237 loci attaining stringent genome-wide significance (P < 5 × 10-9), which were delineated to 338 distinct association signals. Fine-mapping of these signals was enhanced by the increased sample size and expanded population diversity of the multi-ancestry meta-analysis, which localized 54.4% of T2D associations to a single variant with >50% posterior probability. This improved fine-mapping enabled systematic assessment of candidate causal genes and molecular mechanisms through which T2D associations are mediated, laying the foundations for functional investigations. Multi-ancestry genetic risk scores enhanced transferability of T2D prediction across diverse populations. Our study provides a step toward more effective clinical translation of T2D GWAS to improve global health for all, irrespective of genetic background.
Assuntos
Diabetes Mellitus Tipo 2 , Estudo de Associação Genômica Ampla , Diabetes Mellitus Tipo 2/epidemiologia , Etnicidade , Predisposição Genética para Doença , Humanos , Polimorfismo de Nucleotídeo Único/genética , Fatores de RiscoRESUMO
Genome-wide association studies rely on the statistical inference of untyped variants, called imputation, to increase the coverage of genotyping arrays. However, the results are often suboptimal in populations underrepresented in existing reference panels and array designs, since the selected single nucleotide polymorphisms (SNPs) may fail to capture population-specific haplotype structures, hence the full extent of common genetic variation. Here, we propose to sequence the full genomes of a small subset of an underrepresented study cohort to inform the selection of population-specific add-on tag SNPs and to generate an internal population-specific imputation reference panel, such that the remaining array-genotyped cohort could be more accurately imputed. Using a Tanzania-based cohort as a proof-of-concept, we demonstrate the validity of our approach by showing improvements in imputation accuracy after the addition of our designed add-on tags to the base H3Africa array.
Assuntos
Genética Populacional , Estudo de Associação Genômica Ampla , Genótipo , Polimorfismo de Nucleotídeo Único/genética , Biologia Computacional/métodos , Genética Populacional/métodos , Genética Populacional/normas , Estudo de Associação Genômica Ampla/métodos , Estudo de Associação Genômica Ampla/normas , Humanos , Masculino , TanzâniaRESUMO
Trait-associated genetic variants affect complex phenotypes primarily via regulatory mechanisms on the transcriptome. To investigate the genetics of gene expression, we performed cis- and trans-expression quantitative trait locus (eQTL) analyses using blood-derived expression from 31,684 individuals through the eQTLGen Consortium. We detected cis-eQTL for 88% of genes, and these were replicable in numerous tissues. Distal trans-eQTL (detected for 37% of 10,317 trait-associated variants tested) showed lower replication rates, partially due to low replication power and confounding by cell type composition. However, replication analyses in single-cell RNA-seq data prioritized intracellular trans-eQTL. Trans-eQTL exerted their effects via several mechanisms, primarily through regulation by transcription factors. Expression of 13% of the genes correlated with polygenic scores for 1,263 phenotypes, pinpointing potential drivers for those traits. In summary, this work represents a large eQTL resource, and its results serve as a starting point for in-depth interpretation of complex phenotypes.
Assuntos
Proteínas Sanguíneas/genética , Regulação da Expressão Gênica/genética , Locos de Características Quantitativas/genética , Estudo de Associação Genômica Ampla , Humanos , Herança Multifatorial/genética , Polimorfismo de Nucleotídeo Único/genética , Transcriptoma/genéticaRESUMO
Genome-wide association studies (GWAS) are not fully comprehensive, as current strategies typically test only the additive model, exclude the X chromosome, and use only one reference panel for genotype imputation. We implement an extensive GWAS strategy, GUIDANCE, which improves genotype imputation by using multiple reference panels and includes the analysis of the X chromosome and non-additive models to test for association. We apply this methodology to 62,281 subjects across 22 age-related diseases and identify 94 genome-wide associated loci, including 26 previously unreported. Moreover, we observe that 27.7% of the 94 loci are missed if we use standard imputation strategies with a single reference panel, such as HRC, and only test the additive model. Among the new findings, we identify three novel low-frequency recessive variants with odds ratios larger than 4, which need at least a three-fold larger sample size to be detected under the additive model. This study highlights the benefits of applying innovative strategies to better uncover the genetic architecture of complex diseases.
Assuntos
Envelhecimento , Doença/genética , Predisposição Genética para Doença/genética , Genoma Humano/genética , Estudo de Associação Genômica Ampla/métodos , Fatores Etários , Frequência do Gene , Estudo de Associação Genômica Ampla/estatística & dados numéricos , Genótipo , Haplótipos , Humanos , Fenótipo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Epstein-Barr virus (EBV) is one of the most common viruses latently infecting humans. Little is known about the impact of human genetic variation on the large inter-individual differences observed in response to EBV infection. To search for a potential imprint of host genomic variation on the EBV sequence, we jointly analyzed paired viral and human genomic data from 268 HIV-coinfected individuals with CD4 + T cell count < 200/mm3 and elevated EBV viremia. We hypothesized that the reactivated virus circulating in these patients could carry sequence variants acquired during primary EBV infection, thereby providing a snapshot of early adaptation to the pressure exerted on EBV by the individual immune response. We searched for associations between host and pathogen genetic variants, taking into account human and EBV population structure. Our analyses revealed significant associations between human and EBV sequence variation. Three polymorphic regions in the human genome were found to be associated with EBV variation: one at the amino acid level (BRLF1:p.Lys316Glu); and two at the gene level (burden testing of rare variants in BALF5 and BBRF1). Our findings confirm that jointly analyzing host and pathogen genomes can identify sites of genomic interactions, which could help dissect pathogenic mechanisms and suggest new therapeutic avenues.
Assuntos
Variação Genética , Genoma Viral , Herpesvirus Humano 4/genética , Estudos de Coortes , Infecções por Vírus Epstein-Barr/virologia , Sequenciamento de Nucleotídeos em Larga Escala , HumanosRESUMO
Genome-wide association studies (GWAS) have identified thousands of variants associated with complex traits, but their biological interpretation often remains unclear. Most of these variants overlap with expression QTLs, indicating their potential involvement in regulation of gene expression. Here, we propose a transcriptome-wide summary statistics-based Mendelian Randomization approach (TWMR) that uses multiple SNPs as instruments and multiple gene expression traits as exposures, simultaneously. Applied to 43 human phenotypes, it uncovers 3,913 putatively causal gene-trait associations, 36% of which have no genome-wide significant SNP nearby in previous GWAS. Using independent association summary statistics, we find that the majority of these loci were missed by GWAS due to power issues. Noteworthy among these links is educational attainment-associated BSCL2, known to carry mutations leading to a Mendelian form of encephalopathy. We also find pleiotropic causal effects suggestive of mechanistic connections. TWMR better accounts for pleiotropy and has the potential to identify biological mechanisms underlying complex traits.
Assuntos
Estudo de Associação Genômica Ampla , Locos de Características Quantitativas , Encefalopatias/genética , Subunidades gama da Proteína de Ligação ao GTP , Perfilação da Expressão Gênica , Predisposição Genética para Doença , Variação Genética , Humanos , Análise da Randomização Mendeliana , Fenótipo , Polimorfismo de Nucleotídeo Único , TranscriptomaRESUMO
As most of the heritability of complex traits is attributed to common and low frequency genetic variants, imputing them by combining genotyping chips and large sequenced reference panels is the most cost-effective approach to discover the genetic basis of these traits. Association summary statistics from genome-wide meta-analyses are available for hundreds of traits. Updating these to ever-increasing reference panels is very cumbersome as it requires reimputation of the genetic data, rerunning the association scan, and meta-analysing the results. A much more efficient method is to directly impute the summary statistics, termed as summary statistics imputation, which we improved to accommodate variable sample size across SNVs. Its performance relative to genotype imputation and practical utility has not yet been fully investigated. To this end, we compared the two approaches on real (genotyped and imputed) data from 120K samples from the UK Biobank and show that, genotype imputation boasts a 3- to 5-fold lower root-mean-square error, and better distinguishes true associations from null ones: We observed the largest differences in power for variants with low minor allele frequency and low imputation quality. For fixed false positive rates of 0.001, 0.01, 0.05, using summary statistics imputation yielded a decrease in statistical power by 9, 43 and 35%, respectively. To test its capacity to discover novel associations, we applied summary statistics imputation to the GIANT height meta-analysis summary statistics covering HapMap variants, and identified 34 novel loci, 19 of which replicated using data in the UK Biobank. Additionally, we successfully replicated 55 out of the 111 variants published in an exome chip study. Our study demonstrates that summary statistics imputation is a very efficient and cost-effective way to identify and fine-map trait-associated loci. Moreover, the ability to impute summary statistics is important for follow-up analyses, such as Mendelian randomisation or LD-score regression.
Assuntos
Algoritmos , Bioestatística/métodos , Estudo de Associação Genômica Ampla/métodos , Polimorfismo de Nucleotídeo Único , Exoma/genética , Frequência do Gene , Estudos de Associação Genética/métodos , Genótipo , Humanos , Desequilíbrio de Ligação , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Locos de Características Quantitativas/genética , Reprodutibilidade dos TestesRESUMO
There are few examples of robust associations between rare copy number variants (CNVs) and complex continuous human traits. Here we present a large-scale CNV association meta-analysis on anthropometric traits in up to 191,161 adult samples from 26 cohorts. The study reveals five CNV associations at 1q21.1, 3q29, 7q11.23, 11p14.2, and 18q21.32 and confirms two known loci at 16p11.2 and 22q11.21, implicating at least one anthropometric trait. The discovered CNVs are recurrent and rare (0.01-0.2%), with large effects on height (>2.4 cm), weight (>5 kg), and body mass index (BMI) (>3.5 kg/m2). Burden analysis shows a 0.41 cm decrease in height, a 0.003 increase in waist-to-hip ratio and increase in BMI by 0.14 kg/m2 for each Mb of total deletion burden (P = 2.5 × 10-10, 6.0 × 10-5, and 2.9 × 10-3). Our study provides evidence that the same genes (e.g., MC4R, FIBIN, and FMO5) harbor both common and rare variants affecting body size and that anthropometric traits share genetic loci with developmental and psychiatric disorders.Individual SNPs have small effects on anthropometric traits, yet the impact of CNVs has remained largely unknown. Here, Kutalik and co-workers perform a large-scale genome-wide meta-analysis of structural variation and find rare CNVs associated with height, weight and BMI with large effect sizes.
Assuntos
Estatura/genética , Peso Corporal/genética , População Branca/genética , Antropometria , Índice de Massa Corporal , Tamanho Corporal/genética , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 11/genética , Cromossomos Humanos Par 16/genética , Cromossomos Humanos Par 18/genética , Cromossomos Humanos Par 22/genética , Cromossomos Humanos Par 3/genética , Cromossomos Humanos Par 7/genética , Variações do Número de Cópias de DNA , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Fenótipo , Relação Cintura-QuadrilRESUMO
The enormous variation in human lifespan is in part due to a myriad of sequence variants, only a few of which have been revealed to date. Since many life-shortening events are related to diseases, we developed a Mendelian randomization-based method combining 58 disease-related GWA studies to derive longevity priors for all HapMap SNPs. A Bayesian association scan, informed by these priors, for parental age of death in the UK Biobank study (n=116,279) revealed 16 independent SNPs with significant Bayes factor at a 5% false discovery rate (FDR). Eleven of them replicate (5% FDR) in five independent longevity studies combined; all but three are depleted of the life-shortening alleles in older Biobank participants. Further analysis revealed that brain expression levels of nearby genes (RBM6, SULT1A1 and CHRNA5) might be causally implicated in longevity. Gene expression and caloric restriction experiments in model organisms confirm the conserved role for RBM6 and SULT1A1 in modulating lifespan.
Assuntos
Longevidade/genética , Polimorfismo de Nucleotídeo Único , Idoso , Idoso de 80 Anos ou mais , Arilsulfotransferase/genética , Teorema de Bayes , Biomarcadores/análise , Doença/genética , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas de Ligação a RNA/genética , Receptores Nicotínicos/genética , Reino Unido , População Branca/genéticaRESUMO
While hepatitis C exemplifies the role of host genetics in infectious diseases outcomes, there is no comprehensive overview of polymorphisms influencing spontaneous and/or treatment-induced hepatitis C virus clearance. We performed a systematic review and meta-analysis of host polymorphisms associated with these phenotypes. Literature search was conducted using combinations of keywords in three databases. Studies were reviewed and relevant data systematically extracted for subsequent meta-analyses. Polymorphisms from candidate gene studies were tested in two cohorts of HCV-infected patients with available genomic data. The literature search yielded 8'294 citations, among which 262 studies were selected. In the meta-analysis of 27 HLA studies, the most significant associations with spontaneous hepatitis C virus clearance included DQB1*02, DQB1*03, DRB1*04 and DRB1*11. In the meta-analysis of 16 studies of KIR genes and their HLA-ligands, KIR2DS3 was associated with both spontaneous and treatment-induced clearance, and the HLA-C2 ligand with failure to spontaneously clear the virus. In a pooled analysis of 105 candidate genes and two genome-wide association studies, we observed associations of single nucleotide polymorphisms from nine genes (EIF2AK2, IFNAR2, ITPA, MBL2, MX1, OASL, SPP1, TGFB1, TNK2) with response to interferon-based therapy. Meta-analysis of 141 studies confirmed the association of IFNL3/4 polymorphisms with spontaneous and treatment-induced hepatitis C virus clearance, even in previously underpowered groups, such as hepatitis C virus genotypes 2/3-infected patients. This study may contribute to a better understanding of hepatitis C virus immunopathogenesis and highlights the complex role of host genetics in hepatitis C virus clearance.
Assuntos
Antígenos HLA/genética , Hepacivirus/patogenicidade , Hepatite C/genética , Hepatite C/virologia , Polimorfismo de Nucleotídeo Único , Receptores KIR/genética , Antivirais/uso terapêutico , Genótipo , Hepacivirus/efeitos dos fármacos , Hepacivirus/genética , Hepatite C/diagnóstico , Hepatite C/tratamento farmacológico , Interações Hospedeiro-Patógeno , Humanos , Razão de Chances , Fenótipo , Resposta Viral Sustentada , Resultado do TratamentoRESUMO
Height is a highly heritable, classic polygenic trait with approximately 700 common associated variants identified through genome-wide association studies so far. Here, we report 83 height-associated coding variants with lower minor-allele frequencies (in the range of 0.1-4.8%) and effects of up to 2 centimetres per allele (such as those in IHH, STC2, AR and CRISPLD2), greater than ten times the average effect of common variants. In functional follow-up studies, rare height-increasing alleles of STC2 (giving an increase of 1-2 centimetres per allele) compromised proteolytic inhibition of PAPP-A and increased cleavage of IGFBP-4 in vitro, resulting in higher bioavailability of insulin-like growth factors. These 83 height-associated variants overlap genes that are mutated in monogenic growth disorders and highlight new biological candidates (such as ADAMTS3, IL11RA and NOX4) and pathways (such as proteoglycan and glycosaminoglycan synthesis) involved in growth. Our results demonstrate that sufficiently large sample sizes can uncover rare and low-frequency variants of moderate-to-large effect associated with polygenic human phenotypes, and that these variants implicate relevant genes and pathways.
Assuntos
Estatura/genética , Frequência do Gene/genética , Variação Genética/genética , Proteínas ADAMTS/genética , Adulto , Alelos , Moléculas de Adesão Celular/genética , Feminino , Genoma Humano/genética , Glicoproteínas/genética , Glicoproteínas/metabolismo , Glicosaminoglicanos/biossíntese , Proteínas Hedgehog/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Fatores Reguladores de Interferon/genética , Subunidade alfa de Receptor de Interleucina-11/genética , Masculino , Herança Multifatorial/genética , NADPH Oxidase 4 , NADPH Oxidases/genética , Fenótipo , Proteína Plasmática A Associada à Gravidez/metabolismo , Pró-Colágeno N-Endopeptidase/genética , Proteoglicanas/biossíntese , Proteólise , Receptores Androgênicos/genética , Somatomedinas/metabolismoRESUMO
[This corrects the article DOI: 10.1371/journal.pgen.1005378.].
RESUMO
Genome-wide association studies (GWAS) have identified more than 100 genetic variants contributing to BMI, a measure of body size, or waist-to-hip ratio (adjusted for BMI, WHRadjBMI), a measure of body shape. Body size and shape change as people grow older and these changes differ substantially between men and women. To systematically screen for age- and/or sex-specific effects of genetic variants on BMI and WHRadjBMI, we performed meta-analyses of 114 studies (up to 320,485 individuals of European descent) with genome-wide chip and/or Metabochip data by the Genetic Investigation of Anthropometric Traits (GIANT) Consortium. Each study tested the association of up to ~2.8M SNPs with BMI and WHRadjBMI in four strata (men ≤50y, men >50y, women ≤50y, women >50y) and summary statistics were combined in stratum-specific meta-analyses. We then screened for variants that showed age-specific effects (G x AGE), sex-specific effects (G x SEX) or age-specific effects that differed between men and women (G x AGE x SEX). For BMI, we identified 15 loci (11 previously established for main effects, four novel) that showed significant (FDR<5%) age-specific effects, of which 11 had larger effects in younger (<50y) than in older adults (≥50y). No sex-dependent effects were identified for BMI. For WHRadjBMI, we identified 44 loci (27 previously established for main effects, 17 novel) with sex-specific effects, of which 28 showed larger effects in women than in men, five showed larger effects in men than in women, and 11 showed opposite effects between sexes. No age-dependent effects were identified for WHRadjBMI. This is the first genome-wide interaction meta-analysis to report convincing evidence of age-dependent genetic effects on BMI. In addition, we confirm the sex-specificity of genetic effects on WHRadjBMI. These results may provide further insights into the biology that underlies weight change with age or the sexually dimorphism of body shape.
Assuntos
Índice de Massa Corporal , Tamanho Corporal/genética , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Adulto , Fatores Etários , Idoso , Mapeamento Cromossômico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Caracteres Sexuais , Relação Cintura-Quadril , População BrancaRESUMO
Donor PTX3 polymorphisms were shown to influence the risk of invasive aspergillosis among hematopoietic stem cell transplant recipients. Here, we show that PTX3 polymorphisms are independent risk factors for invasive mold infections among 1101 solid organ transplant recipients, thereby strengthening their role in mold infection pathogenesis and patients' risk stratification.