Trajectories of Occupational Cognitive Demands and Risk of Mild Cognitive Impairment and Dementia in Later Life: The HUNT4 70+ Study.

BACKGROUND AND OBJECTIVES: The cognitive reserve hypothesis posits that cognitively stimulating work delays the onset of mild cognitive impairment (MCI) and dementia. However, the effect of occupational cognitive demands across midlife on the risk of these conditions is unclear. METHODS: Using a cohort study design, we evaluated the association between registry-based trajectories of occupational cognitive demands from ages 30-65 years and clinically diagnosed MCI and dementia in participants in the HUNT4 70+ Study (2017-19). Group-based trajectory modeling identified trajectories of occupational cognitive demands, measured by the routine task intensity (RTI) index (lower RTI indicates more cognitively demanding occupation) from the Occupational Information Network. Multinomial regression was implemented to estimate the relative risk ratios (RRRs) of MCI and dementia, after adjusting for age, sex, education, income, baseline hypertension, obesity, diabetes, psychiatric impairment, hearing impairment, loneliness, smoking status, and physical inactivity assessed at HUNT1-2 in 1984-1986 and 1995-1997. To handle missing data, we used inverse probability weighting to account for nonparticipation in cognitive testing and multiple imputation. RESULTS: Based on longitudinal RTI scores for 305 unique occupations, 4 RTI trajectory groups were identified (n = 7,003, 49.8% women, age range 69-104 years): low RTI (n = 1,431, 20.4%), intermediate-low RTI (n = 1,578, 22.5%), intermediate-high RTI (n = 2,601, 37.1%), and high RTI (n = 1,393, 19.9%). Participants in the high RTI group had a higher risk of MCI (RRR 1.74, 95% CI 1.41-2.14) and dementia (RRR 1.37, 95% CI 1.01-1.86), after adjusting for age, sex, and education compared with participants in the low RTI group. In a sensitivity analysis, controlling for income and baseline health-related factors, the point estimates were not appreciably changed (RRR 1.66, 95% CI 1.35-2.06 for MCI, and RRR 1.31, 95% CI 0.96-1.78 for dementia). DISCUSSION: People with a history of cognitively stimulating occupations during their 30s, 40s, 50s, and 60s had a lower risk of MCI and dementia older than 70 years, highlighting the importance of occupational cognitive stimulation during midlife for maintaining cognitive function in old age. Further research is required to pinpoint the specific occupational cognitive demands that are most advantageous for maintaining later-life cognitive function.

Trajectories of occupational physical activity and risk of later-life mild cognitive impairment and dementia: the HUNT4 70+ study.

Background: High levels of occupational physical activity (PA) have been linked to an increased risk of dementia. We assessed the association of trajectories of occupational PA at ages 33-65 with risk of dementia and mild cognitive impairment (MCI) at ages 70+. Methods: We included 7005 participants (49.8% were women, 3488/7005) from the HUNT4 70+ Study. Group-based trajectory modelling was used to identify four trajectories of occupational PA based on national registry data from 1960 to 2014: stable low (30.9%, 2162/7005), increasing then decreasing (8.9%, 625/7005), stable intermediate (25.1%, 1755/7005), and stable high (35.2%, 2463/7005). Dementia and MCI were clinically assessed in 2017-2019. We performed adjusted multinomial regression to estimate relative risk ratios (RRR) with 95% confidence intervals (CI) for dementia and MCI. Findings: 902 participants were diagnosed with dementia and 2407 were diagnosed with MCI. Absolute unadjusted risks for dementia and MCI were 8.8% (95% CI: 7.6-10.0) and 27.4% (25.5-29.3), respectively, for those with a stable low PA trajectory, 8.2% (6.0-10.4) and 33.3% (29.6-37.0) for those with increasing, then decreasing PA; while they were 16.0% (14.3-17.7) and 35% (32.8-37.2) for those with stable intermediate, and 15.4% (14.0-16.8) and 40.2% (38.3-42.1) for those with stable high PA trajectories. In the adjusted model, participants with a stable high trajectory had a higher risk of dementia (RRR 1.34, 1.04-1.73) and MCI (1.80, 1.54-2.11), whereas participants with a stable intermediate trajectory had a higher risk of MCI (1.36, 1.15-1.61) compared to the stable low trajectory. While not statistically significant, participants with increasing then decreasing occupational PA had a 24% lower risk of dementia and 18% higher risk of MCI than the stable low PA group. Interpretation: Consistently working in an occupation with intermediate or high occupational PA was linked to an increased risk of cognitive impairment, indicating the importance of developing strategies for individuals in physically demanding occupations to prevent cognitive impairment. Funding: This work was supported by the National Institutes of Health (R01AG069109-01) and the Research Council of Norway (296297, 262700, 288083).

Statistical methods to detect mother-father genetic interaction effects on risk of infertility: A genome-wide approach.

Infertility is a heterogeneous phenotype, and for many couples, the causes of fertility problems remain unknown. One understudied hypothesis is that allelic interactions between the genotypes of the two parents may influence the risk of infertility. Our aim was, therefore, to investigate how allelic interactions can be modeled using parental genotype data linked to 15,789 pregnancies selected from the Norwegian Mother, Father, and Child Cohort Study. The newborns in 1304 of these pregnancies were conceived using assisted reproductive technologies (ART), and the remainder were conceived naturally. Treating the use of ART as a proxy for infertility, different parameterizations were implemented in a genome-wide screen for interaction effects between maternal and paternal alleles at the same locus. Some of the models were more similar in the way they were parameterized, and some produced similar results when implemented on a genome-wide scale. The results showed near-significant interaction effects in genes relevant to the phenotype under study, such as Dynein axonemal heavy chain 17 (DNAH17) with a recognized role in male infertility. More generally, the interaction models presented here are readily adaptable to the study of other phenotypes in which maternal and paternal allelic interactions are likely to be involved.

Stability selection enhances feature selection and enables accurate prediction of gestational age using only five DNA methylation sites.

BACKGROUND: DNA methylation (DNAm) is robustly associated with chronological age in children and adults, and gestational age (GA) in newborns. This property has enabled the development of several epigenetic clocks that can accurately predict chronological age and GA. However, the lack of overlap in predictive CpGs across different epigenetic clocks remains elusive. Our main aim was therefore to identify and characterize CpGs that are stably predictive of GA. RESULTS: We applied a statistical approach called 'stability selection' to DNAm data from 2138 newborns in the Norwegian Mother, Father, and Child Cohort study. Stability selection combines subsampling with variable selection to restrict the number of false discoveries in the set of selected variables. Twenty-four CpGs were identified as being stably predictive of GA. Intriguingly, only up to 10% of the CpGs in previous GA clocks were found to be stably selected. Based on these results, we used generalized additive model regression to develop a new GA clock consisting of only five CpGs, which showed a similar predictive performance as previous GA clocks (R2 = 0.674, median absolute deviation = 4.4 days). These CpGs were in or near genes and regulatory regions involved in immune responses, metabolism, and developmental processes. Furthermore, accounting for nonlinear associations improved prediction performance in preterm newborns. CONCLUSION: We present a methodological framework for feature selection that is broadly applicable to any trait that can be predicted from DNAm data. We demonstrate its utility by identifying CpGs that are highly predictive of GA and present a new and highly performant GA clock based on only five CpGs that is more amenable to a clinical setting.

Retirement age and disability status as pathways to later-life cognitive impairment: Evidence from the Norwegian HUNT Study linked with Norwegian population registers.

BACKGROUND: Research shows that retirement age is associated with later-life cognition but has not sufficiently distinguished between retirement pathways. We examined how retirement age was associated with later-life dementia and mild cognitive impairment (MCI) for people who retired via the disability pathway (received a disability pension prior to old-age pension eligibility) and those who retired via the standard pathway. METHODS: The study sample comprised 7210 participants from the Norwegian Trøndelag Health Study (HUNT4 70+, 2017-2019) who had worked for at least one year in 1967-2019, worked until age 55+, and retired before HUNT4. Dementia and MCI were clinically assessed in HUNT4 70+ when participants were aged 69-85 years. Historical data on participants' retirement age and pathway were retrieved from population registers. We used multinomial regression to assess the dementia/MCI risk for women and men retiring via the disability pathway, or early (<67 years), on-time (age 67, old-age pension eligibility) or late (age 68+) via the standard pathway. RESULTS: In our study sample, 9.5% had dementia, 35.3% had MCI, and 28.1% retired via the disability pathway. The disability retirement group had an elevated risk of dementia compared to the on-time standard retirement group (relative risk ratio [RRR]: 1.64, 95% CI 1.14-2.37 for women, 1.70, 95% CI 1.17-2.48 for men). MCI risk was lower among men who retired late versus on-time (RRR, 0.76, 95% CI 0.61-0.95). CONCLUSION: Disability retirees should be monitored more closely, and preventive policies should be considered to minimize the dementia risk observed among this group of retirees.

Levels of per- and polyfluoroalkyl substances (PFAS) in Norwegian children stratified by age and sex - Data from the Bergen Growth Study 2.

BACKGROUND AND AIM: Due to the persistence, bioaccumulation and potential adverse health effects, there have been restrictions and phase out in the production of certain per- and polyfluoroalkyl substances (PFAS) since the early 2000s. Published serum levels of PFAS during childhood are variable and may reflect the impact of age, sex, sampling year and exposure history. Surveying the concentrations of PFAS in children is vital to provide information regarding exposure during this critical time of development. The aim of the current study was therefore to evaluate serum concentrations of PFAS in Norwegian schoolchildren according to age and sex. MATERIAL AND METHODS: Serum samples from 1094 children (645 girls and 449 boys) aged 6-16 years, attending schools in Bergen, Norway, were analyzed for 19 PFAS. The samples were collected in 2016 as part of the Bergen Growth Study 2. Statistical analyses included Student t-test, one-way ANOVA and Spearman's correlation analysis of log-transformed data. RESULTS: Of the 19 PFAS examined, 11 were detected in the serum samples. Perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorohexanesulfonic acid (PFHxS) and perfluorononaoic acid (PFNA) were present in all samples with geometric means of 2.67, 1.35, 0.47 and 0.68 ng/mL, respectively. In total, 203 children (19%) had PFAS levels above the safety limits set by the German Human Biomonitoring Commission. Significantly higher serum concentrations were found in boys compared to girls for PFOS, PFNA, PFHxS and perfluoroheptanesulfonic acid (PFHpS). Furthermore, serum concentrations of PFOS, PFOA, PFHxS and PFHpS were significantly higher in children under the age of 12 years than in older children. CONCLUSIONS: PFAS exposure was widespread in the sample population of Norwegian children analyzed in this study. Approximately one out of five children had PFAS levels above safety limits, indicating a potential risk of negative health effects. The majority of the analyzed PFAS showed higher levels in boys than in girls and decreased serum concentrations with age, which may be explained by changes related to growth and maturation.

Deep phenotyping of pubertal development in Norwegian children: the Bergen Growth Study 2.

BACKGROUND: The Bergen Growth Study 2 (BGS2) aims to characterise somatic and endocrine changes in healthy Norwegian children using a novel methodology. SUBJECTS AND METHODS: A cross-sectional sample of 1285 children aged 6-16 years was examined in 2016 using novel objective ultrasound assessments of breast developmental stages and testicular volume in addition to the traditional Tanner pubertal stages. Blood samples allowed for measurements of pubertal hormones, endocrine disruptive chemicals, and genetic analyses. RESULTS: Ultrasound staging of breast development in girls showed a high degree of agreement within and between observers, and ultrasound measurement of testicular volume in boys also showed small intra- and interobserver differences. The median age was 10.4 years for Tanner B2 (pubertal onset) and 12.7 years for menarche. Norwegian boys reached a pubertal testicular volume at a mean age of 11.7 years. Continuous reference curves for testicular volume and sex hormones were constructed using the LMS method. CONCLUSIONS: Ultrasound-based assessments of puberty provided novel references for breast developmental stages and enabled the measurement of testicular volume on a continuous scale. Endocrine z-scores allowed for an intuitive interpretation of changing hormonal levels during puberty on a quantitative scale, which, in turn, provides opportunities for further analysis of pubertal development using machine-learning approaches.

The X-factor in ART: does the use of assisted reproductive technologies influence DNA methylation on the X chromosome?

BACKGROUND: Assisted reproductive technologies (ART) may perturb DNA methylation (DNAm) in early embryonic development. Although a handful of epigenome-wide association studies of ART have been published, none have investigated CpGs on the X chromosome. To bridge this knowledge gap, we leveraged one of the largest collections of mother-father-newborn trios of ART and non-ART (natural) conceptions to date to investigate sex-specific DNAm differences on the X chromosome. The discovery cohort consisted of 982 ART and 963 non-ART trios from the Norwegian Mother, Father, and Child Cohort Study (MoBa). To verify our results from the MoBa cohort, we used an external cohort of 149 ART and 58 non-ART neonates from the Australian 'Clinical review of the Health of adults conceived following Assisted Reproductive Technologies' (CHART) study. The Illumina EPIC array was used to measure DNAm in both datasets. In the MoBa cohort, we performed a set of X-chromosome-wide association studies ('XWASs' hereafter) to search for sex-specific DNAm differences between ART and non-ART newborns. We tested several models to investigate the influence of various confounders, including parental DNAm. We also searched for differentially methylated regions (DMRs) and regions of co-methylation flanking the most significant CpGs. Additionally, we ran an analogous model to our main model on the external CHART dataset. RESULTS: In the MoBa cohort, we found more differentially methylated CpGs and DMRs in girls than boys. Most of the associations persisted after controlling for parental DNAm and other confounders. Many of the significant CpGs and DMRs were in gene-promoter regions, and several of the genes linked to these CpGs are expressed in tissues relevant for both ART and sex (testis, placenta, and fallopian tube). We found no support for parental DNAm-dependent features as an explanation for the observed associations in the newborns. The most significant CpG in the boys-only analysis was in UBE2DNL, which is expressed in testes but with unknown function. The most significant CpGs in the girls-only analysis were in EIF2S3 and AMOT. These three loci also displayed differential DNAm in the CHART cohort. CONCLUSIONS: Genes that co-localized with the significant CpGs and DMRs associated with ART are implicated in several key biological processes (e.g., neurodevelopment) and disorders (e.g., intellectual disability and autism). These connections are particularly compelling in light of previous findings indicating that neurodevelopmental outcomes differ in ART-conceived children compared to those naturally conceived.

Nucleated red blood cells explain most of the association between DNA methylation and gestational age.

Determining if specific cell type(s) are responsible for an association between DNA methylation (DNAm) and a given phenotype is important for understanding the biological mechanisms underlying the association. Our EWAS of gestational age (GA) in 953 newborns from the Norwegian MoBa study identified 13,660 CpGs significantly associated with GA (pBonferroni<0.05) after adjustment for cell type composition. When the CellDMC algorithm was applied to explore cell-type specific effects, 2,330 CpGs were significantly associated with GA, mostly in nucleated red blood cells [nRBCs; n = 2,030 (87%)]. Similar patterns were found in another dataset based on a different array and when applying an alternative algorithm to CellDMC called Tensor Composition Analysis (TCA). Our findings point to nRBCs as the main cell type driving the DNAm-GA association, implicating an epigenetic signature of erythropoiesis as a likely mechanism. They also explain the poor correlation observed between epigenetic age clocks for newborns and those for adults.

Phylogeographic history of mitochondrial haplogroup J in Scandinavia.

BACKGROUND: Mitochondrial DNA haplogroup J is the third most frequent haplogroup in modern-day Scandinavia, although it did not originate there. To infer the genetic history of haplogroup J in Scandinavia, we examined worldwide mitogenome sequences using a maximum-likelihood phylogenetic approach. METHODS: Haplogroup J mitogenome sequences were gathered from GenBank (n = 2245) and aligned against the ancestral Reconstructed Sapiens Reference Sequence. We also analyzed haplogroup J Viking Age sequences from the European Nucleotide Archive (n = 54). Genetic distances were estimated from these data and projected onto a maximum likelihood rooted phylogenetic tree to analyze clustering and branching dates. RESULTS: Haplogroup J originated approximately 42.6 kya (95% CI: 30.0-64.7), with several of its earliest branches being found within the Arabian Peninsula and Northern Africa. J1b was found most frequently in the Near East and Arabian Peninsula, while J1c occurred most frequently in Europe. Based on phylogenetic dating, subhaplogroup J1c has its early roots in the Mediterranean and Western Balkans. Otherwise, the majority of the branches found in Scandinavia are younger than those seen elsewhere, indicating that haplogroup J dispersed relatively recently into Northern Europe, most plausibly with Neolithic farmers. CONCLUSIONS: Haplogroup J appeared when Scandinavia was transitioning to agriculture over 6 kya, with J1c being the most common lineage there today. Changes in the distribution of haplogroup J mtDNAs were likely driven by the expansion of farming from West Asia into Southern Europe, followed by a later expansion into Scandinavia, with other J subhaplogroups appearing among Scandinavian groups as early as the Viking Age.

Marital Histories and Associations With Later-Life Dementia and Mild Cognitive Impairment Risk in the HUNT4 70+ Study in Norway.

Objectives: Earlier studies suggest that being married in later life protects against dementia, and that being single in old age increases the risk of dementia. In this study, we examine midlife marital status trajectories and their association with dementia and mild cognitive impairment (MCI) at ages 70 plus using a large population based sample from Norway. Methods: Based on a general population sample linked to population registries (N = 8706), we used multinomial logistic regression to examine the associations between six types of marital trajectories (unmarried, continuously divorced, intermittently divorced, widowed, continuously married, intermittently married) between age 44 and 68 years from national registries and a clinical dementia or a MCI diagnosis after age 70. We estimated relative risk ratios (RRR) and used mediation analyses adjusting for education, number of children, smoking, hypertension, obesity, physical inactivity, diabetes, mental distress, and having no close friends in midlife. Inverse probability weighting and multiple imputations were applied. The population attributable fraction was estimated to assess the potential reduction in dementia cases due to marital histories. Results: Overall, 11.6% of the participants were diagnosed with dementia and 35.3% with MCI. Dementia prevalence was lowest among the continuously married (11.2%). Adjusting for confounders, the risk of dementia was higher for the unmarried (RRR = 1.73; 95% CI: 1.24, 2.40), continuously divorced (RRR = 1.66; 95% CI: 1.14, 2.43), and intermittently divorced (RRR = 1.50; 95% CI: 1.09, 2.06) compared to the continuously married. In general, marital trajectory was less associated with MCI than with dementia. In the counterfactual scenario, where all participants had the same risk of receiving a dementia diagnosis as the continuously married group, there would be 6.0% fewer dementia cases. Discussion: Our data confirm that staying married in midlife is associated with a lower risk of dementia and that divorced people account for a substantial share of dementia cases.

An examination of mediation by DNA methylation on birthweight differences induced by assisted reproductive technologies.

BACKGROUND: Children born after assisted reproductive technologies (ART) differ in birthweight from those naturally conceived. It has been hypothesized that this might be explained by epigenetic mechanisms. We examined whether cord blood DNA methylation mediated the birthweight difference between 890 newborns conceived by ART (764 by fresh embryo transfer and 126 frozen thawed embryo transfer) and 983 naturally conceived newborns from the Norwegian Mother, Father, and Child Cohort Study (MoBa). DNA methylation was measured by the Illumina Infinium MethylationEPIC array. We conducted mediation analyses to assess whether differentially methylated CpGs mediated the differences in birthweight observed between: (1) fresh embryo transfer and natural conception and (2) frozen and fresh embryo transfer. RESULTS: We observed a difference in birthweight between fresh embryo transfer and naturally conceived offspring of - 120 g. 44% (95% confidence interval [CI] 26% to 81%) of this difference in birthweight between fresh embryo transfer and naturally conceived offspring was explained by differences in methylation levels at four CpGs near LOXL1, CDH20, and DRC1. DNA methylation differences at two CpGs near PTGS1 and RASGRP4 jointly mediated 22% (95% CI 8.1% to 50.3%) of the birthweight differences between fresh and frozen embryo transfer. CONCLUSION: Our findings suggest that DNA methylation is an important mechanism in explaining birthweight differences according to the mode of conception. Further research should examine how gene regulation at these loci influences fetal growth.

Cross-fitted instrument: A blueprint for one-sample Mendelian randomization.

Bias from weak instruments may undermine the ability to estimate causal effects in instrumental variable regression (IVR). We present here a new approach to handling weak instrument bias through the application of a new type of instrumental variable coined 'Cross-Fitted Instrument' (CFI). CFI splits the data at random and estimates the impact of the instrument on the exposure in each partition. These estimates are then used to perform an IVR on each partition. We adapt CFI to the Mendelian randomization (MR) setting and term this adaptation 'Cross-Fitting for Mendelian Randomization' (CFMR). We show that, even when using weak instruments, CFMR is, at worst, biased towards the null, which makes it a conservative one-sample MR approach. In particular, CFMR remains conservative even when the two samples used to perform the MR analysis completely overlap, whereas current state-of-the-art approaches (e.g., MR RAPS) display substantial bias in this setting. Another major advantage of CFMR lies in its use of all of the available data to select genetic instruments, which maximizes statistical power, as opposed to traditional two-sample MR where only part of the data is used to select the instrument. Consequently, CFMR is able to enhance statistical power in consortia-led meta-analyses by enabling a conservative one-sample MR to be performed in each cohort prior to a meta-analysis of the results across all the cohorts. In addition, CFMR enables a cross-ethnic MR analysis by accounting for ethnic heterogeneity, which is particularly important in meta-analyses where the participating cohorts may have different ethnicities. To our knowledge, none of the current MR approaches can account for such heterogeneity. Finally, CFMR enables the application of MR to exposures that are either rare or difficult to measure, which would normally preclude their analysis in the regular two-sample MR setting.

EWAS of post-COVID-19 patients shows methylation differences in the immune-response associated gene, IFI44L, three months after COVID-19 infection.

Although substantial progress has been made in managing COVID-19, it is still difficult to predict a patient's prognosis. We explored the epigenetic signatures of COVID-19 in peripheral blood using data from an ongoing prospective observational study of COVID-19 called the Norwegian Corona Cohort Study. A series of EWASs were performed to compare the DNA methylation profiles between COVID-19 cases and controls three months post-infection. We also investigated differences associated with severity and long-COVID. Three CpGs-cg22399236, cg03607951, and cg09829636-were significantly hypomethylated (FDR < 0.05) in COVID-19 positive individuals. cg03607951 is located in IFI44L which is involved in innate response to viral infection and several systemic autoimmune diseases. cg09829636 is located in ANKRD9, a gene implicated in a wide variety of cellular processes, including the degradation of IMPDH2. The link between ANKRD9 and IMPDH2 is striking given that IMPDHs are considered therapeutic targets for COVID-19. Furthermore, gene ontology analyses revealed pathways involved in response to viruses. The lack of significant differences associated with severity and long-COVID may be real or reflect limitations in sample size. Our findings support the involvement of interferon responsive genes in the pathophysiology of COVID-19 and indicate a possible link to systemic autoimmune diseases.

Associations between epigenetic age acceleration and infertility.

STUDY QUESTION: Is the use of ART, a proxy for infertility, associated with epigenetic age acceleration? SUMMARY ANSWER: The epigenetic age acceleration measured by Dunedin Pace of Aging methylation (DunedinPoAm) differed significantly between non-ART and ART mothers. WHAT IS KNOWN ALREADY: Among mothers who used ART, epigenetic age acceleration may be associated with low oocyte yield and poor ovarian response. However, the difference in epigenetic age acceleration between non-ART and ART mothers (or even fathers) has not been examined. STUDY DESIGN, SIZE, DURATION: The Norwegian Mother, Father and Child Cohort Study (MoBa) recruited pregnant women and their partners across Norway at around 18 gestational weeks between 1999 and 2008. Approximately 95 000 mothers, 75 000 fathers and 114 000 children were included. Peripheral blood samples were taken from mothers and fathers at ultrasound appointments or from mothers at childbirth, and umbilical cord blood samples were collected from the newborns at birth. PARTICIPANTS/MATERIALS, SETTING, METHODS: Among the MoBa participants, we selected 1000 couples who conceived by coitus and 894 couples who conceived by IVF (n = 525) or ICSI (n = 369). We measured their DNA methylation (DNAm) levels using the Illumina MethylationEPIC array and calculated epigenetic age acceleration. A linear mixed model was used to examine the differences in five different epigenetic age accelerations between non-ART and ART parents. MAIN RESULTS AND THE ROLE OF CHANCE: We found a significant difference in the epigenetic age acceleration calculated by DunedinPoAm between IVF and non-ART mothers (0.021 years, P-value = 2.89E-06) after adjustment for potential confounders. Further, we detected elevated DunedinPoAm in mothers with tubal factor infertility (0.030 years, P-value = 1.34E-05), ovulation factor (0.023 years, P-value = 0.0018) and unexplained infertility (0.023 years, P-value = 1.39E-04) compared with non-ART mothers. No differences in epigenetic age accelerations between non-ART and ICSI fathers were found. DunedinPoAm also showed stronger associations with smoking, education and parity than the other four epigenetic age accelerations. LIMITATIONS, REASONS FOR CAUTION: We were not able to determine the directionality of the causal pathway between the epigenetic age accelerations and infertility. Since parents' peripheral blood samples were collected after conception, we cannot rule out the possibility that the epigenetic profile of ART mothers was influenced by the ART treatment. Hence, the results should be interpreted with caution, and our results might not be generalizable to non-pregnant women. WIDER IMPLICATIONS OF THE FINDINGS: A plausible biological mechanism behind the reported association is that IVF mothers could be closer to menopause than non-ART mothers. The pace of decline of the ovarian reserve that eventually leads to menopause varies between females yet, in general, accelerates after the age of 30, and some studies show an increased risk of infertility in females with low ovarian reserve. STUDY FUNDING/COMPETING INTEREST(S): This study was partly funded by the Research Council of Norway (Women's fertility, project no. 320656) and through its Centres of Excellence Funding Scheme (project no. 262700). M.C.M. has received funding from the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation programme (grant agreement number 947684). The authors declare no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.

Age and sex effects on DNA methylation sites linked to genes implicated in severe COVID-19 and SARS-CoV-2 host cell entry.

Male sex and advanced age are associated with severe symptoms of COVID-19. Sex and age also exhibit substantial associations with genome-wide DNA methylation (DNAm) differences in humans. Using a random sample of Illumina EPIC-based genome-wide methylomes from peripheral whole blood of 1,976 parents, participating in The Norwegian Mother, Father and Child Cohort Study (MoBa), we explored whether DNAm in genes linked to SARS-CoV-2 host cell entry and to severe COVID-19 were associated with sex and age. This was carried out by testing 1,572 DNAm sites (CpGs) located near 45 genes for associations with age and sex. We found that DNAm in 281 and 231 of 1,572 CpGs were associated (pFDR<0.01) with sex and aging, respectively. CpGs linked to SARS-CoV-2 host cell entry genes were all associated with age and sex, except for the ACE2 receptor gene (located on the X-chromosome), which was only associated with sex (pFDR<0.01). Furthermore, we examined whether 1,487 autosomal CpGs associated with host-cell entry and severe COVID-19 were more or less associated with sex and age than what would be expected from the same number of randomly sampled genome-wide CpGs. We found that the CpGs associated with host-cell entry and severe COVID-19 were not more or less associated with sex (R2 = 0.77, p = 0.09) than the CpGs sampled from random genomic regions; age was actually found to be significantly less so (R2 = 0.36, p = 0.04). Hence, while we found wide-spread associations between sex and age at CpGs linked to genes implicated with SARS-CoV-2 host cell entry and severe COVID-19, the effect from the sum of these CpGs was not stronger than that from randomly sampled CpGs; for age it was significantly less so. These findings could suggest that advanced age and male sex may not be unsurmountable barriers for the SARS-CoV-2 virus to evolve increased infectiousness.

Evolution and dispersal of mitochondrial DNA haplogroup U5 in Northern Europe: insights from an unsupervised learning approach to phylogeography.

BACKGROUND: We combined an unsupervised learning methodology for analyzing mitogenome sequences with maximum likelihood (ML) phylogenetics to make detailed inferences about the evolution and diversification of mitochondrial DNA (mtDNA) haplogroup U5, which appears at high frequencies in northern Europe. METHODS: Haplogroup U5 mitogenome sequences were gathered from GenBank. The hierarchal Bayesian Analysis of Population Structure (hierBAPS) method was used to generate groups of sequences that were then projected onto a rooted maximum likelihood (ML) phylogenetic tree to visualize the pattern of clustering. The haplogroup statuses of the individual sequences were assessed using Haplogrep2. RESULTS: A total of 23 hierBAPS groups were identified, all of which corresponded to subclades defined in Phylotree, v.17. The hierBAPS groups projected onto the ML phylogeny accurately clustered all haplotypes belonging to a specific haplogroup in accordance with Haplogrep2. By incorporating the geographic source of each sequence and subclade age estimates into this framework, inferences about the diversification of U5 mtDNAs were made. Haplogroup U5 has been present in northern Europe since the Mesolithic, and spread in both eastern and western directions, undergoing significant diversification within Scandinavia. A review of historical and archeological evidence attests to some of the population interactions contributing to this pattern. CONCLUSIONS: The hierBAPS algorithm accurately grouped mitogenome sequences into subclades in a phylogenetically robust manner. This analysis provided new insights into the phylogeographic structure of haplogroup U5 diversity in northern Europe, revealing a detailed perspective on the diversity of subclades in this region and their distribution in Scandinavian populations.

DNA methylation in newborns conceived by assisted reproductive technology.

Assisted reproductive technology (ART) may affect fetal development through epigenetic mechanisms as the timing of ART procedures coincides with the extensive epigenetic remodeling occurring between fertilization and embryo implantation. However, it is unknown to what extent ART procedures alter the fetal epigenome. Underlying parental characteristics and subfertility may also play a role. Here we identify differences in cord blood DNA methylation, measured using the Illumina EPIC platform, between 962 ART conceived and 983 naturally conceived singleton newborns. We show that ART conceived newborns display widespread differences in DNA methylation, and overall less methylation across the genome. There were 607 genome-wide differentially methylated CpGs. We find differences in 176 known genes, including genes related to growth, neurodevelopment, and other health outcomes that have been associated with ART. Both fresh and frozen embryo transfer show DNA methylation differences. Associations persist after controlling for parents' DNA methylation, and are not explained by parental subfertility.

Wavelet Screening: a novel approach to analyzing GWAS data.

BACKGROUND: Traditional methods for single-variant genome-wide association study (GWAS) incur a substantial multiple-testing burden because of the need to test for associations with a vast number of single-nucleotide polymorphisms (SNPs) simultaneously. Further, by ignoring more complex joint effects of nearby SNPs within a given region, these methods fail to consider the genomic context of an association with the outcome. RESULTS: To address these shortcomings, we present a more powerful method for GWAS, coined 'Wavelet Screening' (WS), that greatly reduces the number of tests to be performed. This is achieved through the use of a sliding-window approach based on wavelets to sequentially screen the entire genome for associations. Wavelets are oscillatory functions that are useful for analyzing the local frequency and time behavior of signals. The signals can then be divided into different scale components and analyzed separately. In the current setting, we consider a sequence of SNPs as a genetic signal, and for each screened region, we transform the genetic signal into the wavelet space. The null and alternative hypotheses are modeled using the posterior distribution of the wavelet coefficients. WS is enhanced by using additional information from the regression coefficients and by taking advantage of the pyramidal structure of wavelets. When faced with more complex genetic signals than single-SNP associations, we show via simulations that WS provides a substantial gain in power compared to both the traditional GWAS modeling and another popular regional association test called SNP-set (Sequence) Kernel Association Test (SKAT). To demonstrate feasibility, we applied WS to a large Norwegian cohort (N=8006) with genotypes and information available on gestational duration. CONCLUSIONS: WS is a powerful and versatile approach to analyzing whole-genome data and lends itself easily to investigating various omics data types. Given its broader focus on the genomic context of an association, WS may provide additional insight into trait etiology by revealing genes and loci that might have been missed by previous efforts.