Integrating biological knowledge and gene expression data using pathway-guided random forests: a benchmarking study.

MOTIVATION: High-throughput technologies allow comprehensive characterization of individuals on many molecular levels. However, training computational models to predict disease status based on omics data is challenging. A promising solution is the integration of external knowledge about structural and functional relationships into the modeling process. We compared four published random forest-based approaches using two simulation studies and nine experimental datasets. RESULTS: The self-sufficient prediction error approach should be applied when large numbers of relevant pathways are expected. The competing methods hunting and learner of functional enrichment should be used when low numbers of relevant pathways are expected or the most strongly associated pathways are of interest. The hybrid approach synthetic features is not recommended because of its high false discovery rate. AVAILABILITY AND IMPLEMENTATION: An R package providing functions for data analysis and simulation is available at GitHub (https://github.com/szymczak-lab/PathwayGuidedRF). An accompanying R data package (https://github.com/szymczak-lab/DataPathwayGuidedRF) stores the processed and quality controlled experimental datasets downloaded from Gene Expression Omnibus (GEO). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Sex-specific genetic factors affect the risk of early-onset periodontitis in Europeans.

AIMS: Various studies have reported that young European women are more likely to develop early-onset periodontitis compared to men. A potential explanation for the observed variations in sex and age of disease onset is the natural genetic variation within the autosomal genomes. We hypothesized that genotype-by-sex (G × S) interactions contribute to the increased prevalence and severity. MATERIALS AND METHODS: Using the case-only design, we tested for differences in genetic effects between men and women in 896 North-West European early-onset cases, using imputed genotypes from the OmniExpress genotyping array. Population-representative 6823 controls were used to verify that the interacting variables G and S were uncorrelated in the general population. RESULTS: In total, 20 loci indicated G × S associations (P < 0.0005), 3 of which were previously suggested as risk genes for periodontitis (ABLIM2, CDH13, and NELL1). We also found independent G × S interactions of the related gene paralogs MACROD1/FLRT1 (chr11) and MACROD2/FLRT3 (chr20). G × S-associated SNPs at CPEB4, CDH13, MACROD1, and MECOM were genome-wide-associated with heel bone mineral density (CPEB4, MECOM), waist-to-hip ratio (CPEB4, MACROD1), and blood pressure (CPEB4, CDH13). CONCLUSIONS: Our results indicate that natural genetic variation affects the different heritability of periodontitis among sexes and suggest genes that contribute to inter-sex phenotypic variation in early-onset periodontitis.

Rare Variants in Specific Lysosomal Genes Are Associated With Parkinson's Disease.

OBJECTIVE: Impaired lysosomal degradation of α-synuclein and other cellular constituents may play an important role in Parkinson's disease (PD). Rare genetic variants in the glucocerebrosidase (GBA) gene were consistently associated with PD. Here we examine the association between rare variants in lysosomal candidate genes and PD. METHODS: We investigated the association between PD and rare genetic variants in 23 lysosomal candidate genes in 4096 patients with PD and an equal number of controls using pooled targeted next-generation DNA sequencing. Genewise association of rare variants in cases or controls was analyzed using the optimized sequence kernel association test with Bonferroni correction for the 23 tested genes. RESULTS: We confirm the association of rare variants in GBA with PD and report novel associations for rare variants in ATP13A2, LAMP1, TMEM175, and VPS13C. CONCLUSION: Rare variants in selected lysosomal genes, first and foremost GBA, are associated with PD. Rare variants in ATP13A2 and VPC13C previously linked to monogenic PD and more common variants in TMEM175 and VPS13C previously linked to sporadic PD in genome-wide association studies are associated with PD. © 2020 International Parkinson and Movement Disorder Society.

IMHOTEP-a composite score integrating popular tools for predicting the functional consequences of non-synonymous sequence variants.

The in silico prediction of the functional consequences of mutations is an important goal of human pathogenetics. However, bioinformatic tools that classify mutations according to their functionality employ different algorithms so that predictions may vary markedly between tools. We therefore integrated nine popular prediction tools (PolyPhen-2, SNPs&GO, MutPred, SIFT, MutationTaster2, Mutation Assessor and FATHMM as well as conservation-based Grantham Score and PhyloP) into a single predictor. The optimal combination of these tools was selected by means of a wide range of statistical modeling techniques, drawing upon 10 029 disease-causing single nucleotide variants (SNVs) from Human Gene Mutation Database and 10 002 putatively 'benign' non-synonymous SNVs from UCSC. Predictive performance was found to be markedly improved by model-based integration, whilst maximum predictive capability was obtained with either random forest, decision tree or logistic regression analysis. A combination of PolyPhen-2, SNPs&GO, MutPred, MutationTaster2 and FATHMM was found to perform as well as all tools combined. Comparison of our approach with other integrative approaches such as Condel, CoVEC, CAROL, CADD, MetaSVM and MetaLR using an independent validation dataset, revealed the superiority of our newly proposed integrative approach. An online implementation of this approach, IMHOTEP ('Integrating Molecular Heuristics and Other Tools for Effect Prediction'), is provided at http://www.uni-kiel.de/medinfo/cgi-bin/predictor/.

Thinning of Inner Retinal Layers after Vitrectomy with Silicone Oil versus Gas Endotamponade in Eyes with Macula-Off Retinal Detachment.

PURPOSE: To evaluate retinal layer thickness with optical coherence tomography (OCT) in eyes with macula-off retinal detachment after silicone oil (SiO) or gas endotamponade. PROCEDURES: Cross-sectional study of 40 eyes with macula-off rhegmatogenous retinal detachment that underwent vitrectomy. 20 eyes received SiO tamponade and 20 matched eyes received gas. 33 healthy fellow eyes served as controls. Macular spectral domain OCT was performed with automated layer detection in the 5 inner subfields of the Early Treatment Diabetic Retinopathy Study (ETDRS) map. RESULTS: Comparing the SiO group with the gas group, the ganglion cell layer showed a significant thinning in all fields of the inner ring of the ETDRS map, the inner plexiform layer in the nasal, superior and temporal quadrants, and the outer plexiform layer in the nasal quadrant. CONCLUSIONS: Inner retinal layers in the fovea/parafovea were significantly thinner in the SiO group. Prospective studies are warranted to further elucidate possible retinal adverse effects of SiO tamponade.

Network-based quantitative trait linkage analysis of microbiome composition in inflammatory bowel disease families.

Introduction: Inflammatory bowel disease (IBD) is characterized by a dysbiosis of the gut microbiome that results from the interaction of the constituting taxa with one another, and with the host. At the same time, host genetic variation is associated with both IBD risk and microbiome composition. Methods: In the present study, we defined quantitative traits (QTs) from modules identified in microbial co-occurrence networks to measure the inter-individual consistency of microbial abundance and subjected these QTs to a genome-wide quantitative trait locus (QTL) linkage analysis. Results: Four microbial network modules were consistently identified in two cohorts of healthy individuals, but three of the corresponding QTs differed significantly between IBD patients and unaffected individuals. The QTL linkage analysis was performed in a sub-sample of the Kiel IBD family cohort (IBD-KC), an ongoing study of 256 German families comprising 455 IBD patients and 575 first- and second-degree, non-affected relatives. The analysis revealed five chromosomal regions linked to one of three microbial module QTs, namely on chromosomes 3 (spanning 10.79 cM) and 11 (6.69 cM) for the first module, chr9 (0.13 cM) and chr16 (1.20 cM) for the second module, and chr13 (19.98 cM) for the third module. None of these loci have been implicated in a microbial phenotype before. Discussion: Our study illustrates the benefit of combining network and family-based linkage analysis to identify novel genetic drivers of microbiome composition in a specific disease context.

Correlation between genetic and geographic structure in Europe.

Understanding the genetic structure of the European population is important, not only from a historical perspective, but also for the appropriate design and interpretation of genetic epidemiological studies. Previous population genetic analyses with autosomal markers in Europe either had a wide geographic but narrow genomic coverage [1, 2], or vice versa [3-6]. We therefore investigated Affymetrix GeneChip 500K genotype data from 2,514 individuals belonging to 23 different subpopulations, widely spread over Europe. Although we found only a low level of genetic differentiation between subpopulations, the existing differences were characterized by a strong continent-wide correlation between geographic and genetic distance. Furthermore, mean heterozygosity was larger, and mean linkage disequilibrium smaller, in southern as compared to northern Europe. Both parameters clearly showed a clinal distribution that provided evidence for a spatial continuity of genetic diversity in Europe. Our comprehensive genetic data are thus compatible with expectations based upon European population history, including the hypotheses of a south-north expansion and/or a larger effective population size in southern than in northern Europe. By including the widely used CEPH from Utah (CEU) samples into our analysis, we could show that these individuals represent northern and western Europeans reasonably well, thereby confirming their assumed regional ancestry.

Comparison of Markov Chain Monte Carlo Software for the Evolutionary Analysis of Y-Chromosomal Microsatellite Data.

The evolutionary analysis of genetic data is an important subject of modern bioscience, with practical applications in diverse fields. Parameters of interest in this context include effective population sizes, mutation rates, population growth rates and the times to most recent common ancestors. Studying Y-chromosomal microsatellite data, in particular, has proven useful to unravel the recent patrilineal history of Homo sapiens populations. We compared the individual analysis options and technical details of four software tools that are widely used for this purpose, namely BATWING, BEAST, IMa2 and LAMARC, all of which use Bayesian coalescent-based Markov chain Monte Carlo (MCMC) methods for parameter estimation. More specifically, we simulated datasets for either eight or 20 hypothetical Y-chromosomal microsatellites, assuming a mutation rate of 0.0030 per generation and a constant or exponentially increasing population size, and used these data to evaluate the parameter estimation capacity of each tool. The datasets comprised between 100 and 1000 samples. In addition to runtime, the practical utility of the tools of interest can also be expected to depend critically upon the convergence behavior of the actual MCMC implementation. In fact, we found that runtime increased, and convergence rate decreased, with increasing sample size as expected. BATWING performed best with respect to runtime and convergence behavior, but only supports simple evolutionary models. As regards the spectrum of evolutionary models covered, and also in terms of cross-platform usability, BEAST provided the greatest flexibility. Finally, IMa2 and LAMARC turned out best to incorporate elaborate migration models in the analysis process.

VarWatch-A stand-alone software tool for variant matching.

Massively parallel DNA sequencing of clinical samples holds great promise for the gene-based diagnosis of human inherited diseases because it allows rapid detection of putatively causative mutations at genome-wide level. Without additional evidence complementing their initial bioinformatics evaluation, however, the clinical relevance of such candidate genetic variants often remains unclear. In consequence, dedicated 'matching' services have been established in recent years that aim at the discovery of other, comparable case reports to facilitate individual diagnoses. However, legal concerns have been raised about the global sharing of genetic data, particularly in Europe where the recently enacted General Data Protection Regulation EU-2016/679 classifies genetic data as highly sensitive. Hence, unrestricted sharing of genetic data from clinical cases on platforms outside the national jurisdiction increasingly may be perceived as problematic. To allow collaborative data producers, particularly large consortia of diagnostic laboratories, to acknowledge these concerns while still practicing efficient case matching internally, novel tools are required. To this end, we developed VarWatch, an easy-to-deploy and highly scalable case matching software that provides users with comprehensive programmatic tools and a user-friendly interface to fulfil said purpose.

Low incidence of inbreeding in a long-lived primate population isolated for 75 years.

ABSTRACT: When close relatives mate, offspring are expected to suffer fitness consequences due to inbreeding depression. Inbreeding has previously been quantified in two ways: using a sufficiently large panel of markers or deep and complete pedigrees over several generations. However, the application of both approaches is still limited by the challenge of compiling such data for species with long generation times, such as primates. Here, we assess inbreeding in rhesus macaques living on Cayo Santiago (Puerto Rico), a population genetically isolated since 1938, but descendant of a large set of presumably unrelated founders. Using comprehensive genetic data, we calculated inbreeding coefficients (F) for 2669 individuals with complete three generation pedigrees and 609 individuals with complete four generation pedigrees. We found that 0.79 and 7.39% of individuals had an F > 0 when using data from three and four generation pedigrees, respectively. No evidence of an increase in inbreeding over the study period (up to 23 years) was found. Furthermore, the observed mean relatedness of breeding pairs differed significantly from the distribution of parental relatedness expected as simulated based on previous reproductive data, suggesting that kin generally avoid breeding with each other. Finally, inbreeding was not a predictor of early mortality measured as survival until weaning and sexual maturation, respectively. Our results remain consistent with three estimators of inbreeding (standardized heterozygosity, internal relatedness, and homozygosity by loci) using up to 42 highly polymorphic microsatellites for the same set of individuals. Together, our results demonstrate that close inbreeding may not be prevalent even in populations isolated over long periods when mechanisms of inbreeding avoidance can operate. SIGNIFICANCE STATEMENT: When close relatives mate, offspring may suffer from such inbreeding, e.g., via lower survival and/or fertility. Using (i) a large panel of genetic markers and (ii) complete three or four generation pedigrees, respectively, we show that incidences of inbreeding in a long-lived primate population are rare, even after genetic isolation for 75 years. Moreover, our simulations suggest that kin in our population generally avoid breeding with each other. Finally, the few inbred individuals detected in our large sample did not suffer from lower survival. Given that many animal species face dramatic habitat loss combined with critical population declines, our study provides important implications for conservation biology in general and for population management in particular.

Forensic interpretation of Y-chromosomal DNA mixtures.

The mathematical concept previously introduced for the forensic interpretation of DNA mixtures using non-associated genetic markers has been adapted to the assessment of haplotypes. Such calculus is required, for example, when Y-chromosomal markers are used in forensics. In addition to outlining the general mathematical framework, we devise two approaches to its practical computational implementation, involving either the inclusion-exclusion principle of probability theory or a recursion in the number of unknown contributors invoked. The two approaches scale differently, depending upon the complexity of the case and the diversity of the markers used. The performance of Y-chromosomal microsatellites (Y-STRs) as a means of trace donor discrimination has been assessed by simulation, using the derived formulas. Based upon data from the Y-chromosomal Haplotype Reference Database (YHRD), the exclusion chance of a non-contributor is shown to vary between 95% in the case of two contributors, and 70% for five contributors. With only one additional contributor, half of all contributing suspects would yield a log-likelihood ratio in favour of donorship of 1.61 or higher, although the median drops to 0.66 with four additional contributors. It must be emphasised that these estimates of the discriminatory power of Y-STRs are likely to be conservative since the simulations involved only haplotypes known to occur in YHRD.

The long-term course of functional and anatomical recovery after macular hole surgery.

PURPOSE: To investigate the long-term effect of macular hole surgery, foveal structure and the thickness of retinal layers were analyzed with spectral-domain optical coherence tomography (SD-OCT). The long-term postoperative course of macular thickness and best-corrected visual acuity (BCVA) were followed. METHODS: In a retrospective cohort study, SD-OCT scans were obtained from the horizontal midline in 51 eyes 54±20 months postoperatively and from 30 control eyes. Retinal layer thickness was measured with a manual segmentation procedure aided by a customized computer program. BCVA was followed and macular thickness was quantified over time with the time-domain (TD) OCT Fast Macular Thickness program for up to 91 months. RESULTS: Median foveal thickness between the outer plexiform and ganglion cell layers was greater than normal while that of the other retinal layers was normal. The median foveal shape remained slightly distorted. The postoperative decrease of central macular thickness toward normal values was delayed to 28 months postoperatively. Nasal macular thickness was decreased to normal at 6 months while superior, temporal, and inferior macular thickness was decreased to normal at 1 to 2 months postoperatively. Preoperative mean BCVA was 20/100±3 lines. Postoperatively, mean BCVA was 20/44±2 lines at 3 to 6 months, 20/40±2 lines at 1 year, 20/32±2 lines at 2 years, and 20/28±1 line after a mean follow-up period of 54±20 months. CONCLUSIONS: Long-term postoperatively, the median thickness of retinal layers remains slightly thickened between the outer plexiform and the ganglion cell layer. The process of gradual recovery may continue for several years after macular hole surgery.

Foveal structure and thickness of retinal layers long-term after surgical peeling of idiopathic epiretinal membrane.

PURPOSE: To better understand the long-term effect of idiopathic epiretinal membrane peeling on retinal anatomy, the foveal structure and the thickness of individual retinal layers were analyzed with frequency-domain optical coherence tomography (fdOCT). The long-term postoperative course of macular thickness was followed. METHODS: fdOCT scans were obtained from the horizontal midline in 33 eyes long-term (mean 46 ± 13 months) after surgery and in 30 eyes of age-matched controls. Raw images were exported, and the thickness of retinal layers was measured with a manual segmentation procedure aided by a customized computer program. Macular thickness was quantified over time with the time-domain (td) OCT Fast Macular Thickness program. RESULTS: Thickness of retinal layers between the outer nuclear and the ganglion cell plus inner plexiform layers in the horizontal midline of the fovea and the nasal parafovea was greater than normal, whereas that of the RPE, photoreceptor, and retinal nerve fiber layers was not different from controls. Twelve of 33 eyes had a foveal pit though the median foveal shape was distorted. Central macular thickness quantified with tdOCT remained increased, whereas the decrease of nasal macular thickness toward normal values was incomplete and delayed to 35 months after surgery. Superior, temporal, and inferior macular thickness returned to normal 12 to 14 months after surgery. CONCLUSIONS: Long-term after surgery, the fovea and the nasal parafovea remain thickened between the outer nuclear layer and the ganglion cell layer, whereas the superior, temporal, and inferior macular thickness returns to normal. Long-term observations are required in the assessment of macular recovery from mechanical stress.

A genome-wide association study confirms APOE as the major gene influencing survival in long-lived individuals.

We conducted a case-control genome-wide association study (GWAS) of human longevity, comparing 664,472 autosomal SNPs in 763 long-lived individuals (LLI; mean age: 99.7 years) and 1085 controls (mean age: 60.2 years) from Germany. Only one association, namely that of SNP rs4420638 near the APOC1 gene, achieved genome-wide significance (allele-based P=1.8×10(-10)). However, logistic regression analysis revealed that this association, which was replicated in an independent German sample, is fully explicable by linkage disequilibrium with the APOE allele ɛ4, the only variant hitherto established as a major genetic determinant of survival into old age. Our GWAS failed to identify any additional autosomal susceptibility genes. One explanation for this lack of success in our study would be that GWAS provide only limited statistical power for a polygenic phenotype with loci of small effect such as human longevity. A recent GWAS in Dutch LLI independently confirmed the APOE-longevity association, thus strengthening the conclusion that this locus is a very, if not the most, important genetic factor influencing longevity.

An evaluation of the genetic-matched pair study design using genome-wide SNP data from the European population.

Genetic matching potentially provides a means to alleviate the effects of incomplete Mendelian randomization in population-based gene-disease association studies. We therefore evaluated the genetic-matched pair study design on the basis of genome-wide SNP data (309,790 markers; Affymetrix GeneChip Human Mapping 500K Array) from 2457 individuals, sampled at 23 different recruitment sites across Europe. Using pair-wise identity-by-state (IBS) as a matching criterion, we tried to derive a subset of markers that would allow identification of the best overall matching (BOM) partner for a given individual, based on the IBS status for the subset alone. However, our results suggest that, by following this approach, the prediction accuracy is only notably improved by the first 20 markers selected, and increases proportionally to the marker number thereafter. Furthermore, in a considerable proportion of cases (76.0%), the BOM of a given individual, based on the complete marker set, came from a different recruitment site than the individual itself. A second marker set, specifically selected for ancestry sensitivity using singular value decomposition, performed even more poorly and was no more capable of predicting the BOM than randomly chosen subsets. This leads us to conclude that, at least in Europe, the utility of the genetic-matched pair study design depends critically on the availability of comprehensive genotype information for both cases and controls.