Prenatal folate, homocysteine and vitamin B12 levels and child brain volumes, cognitive development and psychological functioning: the Generation R Study.

Previous studies have suggested that prenatal maternal folate deficiency is associated with reduced prenatal brain growth and psychological problems in offspring. However, little is known about the longer-term impact. The aims of this study were to investigate whether prenatal maternal folate insufficiency, high total homocysteine levels and low vitamin B12 levels are associated with altered brain morphology, cognitive and/or psychological problems in school-aged children. This study was embedded in Generation R, a prospective population-based cohort study. The study sample consisted of 256 Dutch children aged between 6 and 8 years from whom structural brain scans were collected using MRI. The mothers of sixty-two children had insufficient (<8 nmol/l) plasma folate concentrations in early pregnancy. Cognitive development was assessed by the Snijders-Oomen Niet-verbale intelligentietest - Revisie and the NEPSY-II-NL. Psychological problems were assessed at age 6 years using the parent report of the Child Behavior Checklist. Low prenatal folate levels were associated with a smaller total brain volume (B -33·34; 95 % CI -66·7, 0·02; P=050) and predicted poorer performance on the language (B -0·28; 95 % CI -0·52, -0·04; P=0·020) and visuo-spatial domains (B -0·27; 95 % CI -0·50, -0·04; P=0·021). High homocysteine levels (>9·1 µmol/l) predicted poorer performance on the language (B -0·31; 95 % CI -0·56, -0·06; P=0·014) and visuo-spatial domains (B -0·36; 95 % CI -0·60, -0·11; P=0·004). No associations with psychological problems were found. Our findings suggest that folate insufficiency in early pregnancy has a long-lasting, global effect on brain development and is, together with homocysteine levels, associated with poorer cognitive performance.

Cultural effects on neurodevelopmental testing in children from six European countries: an analysis of NUTRIMENTHE Global Database.

Cultural background is an important variable influencing neuropsychological performance. Multinational projects usually involve gathering data from participants from different countries and/or different cultures. Little is known about the influence of culture on neuropsychological testing results in children and especially in European children. The objectives of this study were to compare neuropsychological performance of children from six European countries (Belgium, Germany, Italy, The Netherlands, Poland and Spain) using a comprehensive neuropsychological battery and to apply a statistical procedure to reduce the influence of country/cultural differences in neuropsychological performance. As expected, the results demonstrated differences in neuropsychological performance among children of the six countries involved. Cultural differences remained after adjusting for other confounders related to neuropsychological execution, such as sex, type of delivery, maternal age, gestational age and maternal educational level. Differences between countries disappeared and influence of culture was considerably reduced when standardised scores by country and sex were used. These results highlight the need for developing specific procedures to compare neuropsychological performance among children from different cultures to be used in multicentre studies.

Maternal plasma n-3 and n-6 polyunsaturated fatty acids during pregnancy and features of fetal health: Fetal growth velocity, birth weight and duration of pregnancy.

BACKGROUND & AIMS: Maternal fatty acids are essential for fetal growth and development. Here, we examine associations between maternal mid-pregnancy plasma n-3 and n-6 polyunsaturated fatty acids (PUFAs) and fetal health determined by fetal growth velocity, birth weight and duration of pregnancy. METHODS: Participants were 6974 pregnant women and their infants from a population-based birth cohort, the Generation R Study. Maternal plasma n-3:n-6 PUFA ratio and n-3 and n-6 PUFA percentage in glycerophospholipids in mid-pregnancy were related to fetal growth velocity calculated from repeatedly measured weight, length and head circumference, birth weight, and duration of pregnancy. RESULTS: A higher maternal mid-pregnancy n-3:n-6 PUFA ratio was associated with a higher growth velocity of the fetal weight (ß = 0.082 SD-score/week, 95% CI 0.055; 0.108, P < 0.001), length (ß = 0.085 SD-score/week, 95% CI 0.052; 0.119, P < 0.001); and head (ß = 0.055 SD-score/week, 95% CI 0.019; 0.091, P = 0.003). We also observed positive associations between n-3:n-6 PUFA ratio and birth weight (ß = 0.76 SD-score, 95% CI 0.22; 1.29, P = 0.006), and duration of pregnancy (ß = 1.32 weeks, 95% CI 0.24; 2.40, P = 0.02). CONCLUSIONS: These results are consistent with the hypothesis that a higher n-3:n-6 PUFA ratio is important for fetal health.

Plasma fatty acid patterns during pregnancy and child's growth, body composition, and cardiometabolic health: The Generation R Study.

BACKGROUND: Exposure to different concentrations of fatty acids during fetal life may affect growth and metabolism. However, most studies examined individual fatty acids, whereas concentrations highly correlate and may interact with each other. We aimed to evaluate patterns of plasma fatty acids during pregnancy and their associations with growth, body composition, and cardiometabolic health of the 6-year-old offspring. METHODS: This study was performed in 4830 mother-child pairs participating in a population-based cohort in the Netherlands. Around 20 weeks of gestation, we measured plasma phospholipid concentrations of 22 fatty acids, in which we identified three fatty acid patterns using principal component analysis: a 'high n-6 polyunsaturated fatty acid (PUFA)' pattern, a 'monounsaturated and saturated fatty acid (MUFA and SFA)' pattern, and a 'high n-3 PUFA' pattern. When the children were 6 years old, we measured their anthropometrics and detailed body composition (using dual-energy X-ray absorptiometry), and we calculated their body mass index (BMI), fat mass index (FMI), fat-free mass index (FFMI). At the same age, children's blood pressure, and serum insulin, HDL-cholesterol, and triacylglycerol were measured. RESULTS: After adjustment for confounders and the other patterns, a higher score for the 'high n-6 PUFA' pattern during pregnancy was associated with a higher height, BMI, and FFMI in the offspring at 6 years, but not independently with cardiometabolic outcomes. The 'MUFA and SFA' pattern was not consistently associated with child body composition or cardiometabolic health. A higher score for the 'high n-3 PUFA' pattern was associated with a lower FMI, higher FFMI, higher HDL-cholesterol, and lower triacylglycerol. CONCLUSIONS: Our results suggest that plasma fatty acid patterns during pregnancy may affect offspring's body composition and cardiometabolic health. Specifically, a pattern characterized by high n-3 PUFA levels was associated with a more favorable body composition and blood lipid profile.

Infant feeding and child fussy eating: The Generation R Study.

Fussy/picky eating - i.e. consistently avoiding certain foods - is common in childhood and can be worrisome for parents. Repeated exposure to various flavors as occurs in breastmilk and early exposure to complementary feeding may increase food acceptance and thereby decrease fussy eating. This study examines the associations between infant feeding and child fussy eating in 4779 participants of Generation R, a Dutch population-based cohort. Breastfeeding initiation and continuation, and timing of complementary feeding were assessed by questionnaires at 2, 6, and 12 months. The food fussiness scale of the Children's Eating Behaviour Questionnaire was administered at 4 years. Linear regression analyses were performed, adjusted for confounders. Children who were never breastfed did not differ in fussy eating frequency from children breastfed for 6 months or longer. However, children who were breastfed for less than 2 months had a 0.70 points higher food fussiness sum-score (95%CI:0.27; 1.12) than children breastfed for 6 months or longer. An earlier introduction of vegetables was associated with less fussy eating behavior (p-for-trend:0.005). Particularly children who were introduced to vegetables between 4 and 5 months had a 0.60 point lower food fussiness score (95%CI: 1.06;-0.15) than children introduced to vegetables after 6 months. An early introduction to fruits or any solids was not significantly related to fussy eating, although the effect estimates were in the same direction as for introducing vegetables early. Results suggest that breastfeeding does not predict fussy eating. However, introducing vegetables into a child's diet before 5 months might be protective against fussy eating, although future research should account for parents' own fussy eating.

Maternal Midpregnancy Plasma trans 18:1 Fatty Acid Concentrations Are Positively Associated with Risk of Maternal Vascular Complications and Child Low Birth Weight.

Background: Evidence is plentiful that trans fatty acids (TFAs) induce vascular inflammation with adverse metabolic consequences. However, it is not clear whether TFAs increase the risk of vascular pregnancy complications such as preeclampsia.Objective: We investigated associations between midpregnancy maternal plasma trans 18:1 fatty acid (t18:1) concentrations and pregnancy course and outcomes.Methods: Participants were 6695 pregnant women and newborns from the Generation R Study, Rotterdam, Netherlands (enrollment in 2001-2005). Maternal midpregnancy (mean ± SD gestational age: 20.7 ± 1.2 wk) t18:1 plasma concentrations were determined and related to gestational age and sex-adjusted birth weight SD scores, placental weight, and the risk of preeclampsia. In addition, we explored potential time trends by testing the association of maternal plasma t18:1 concentrations with birth weight in birth cohorts given the Dutch industry-initiative to lower food TFA contents during the inclusion period. Multiple logistic and linear regression analyses were performed, taking various socioeconomic and biological covariates into account.Results: A higher midpregnancy maternal plasma t18:1 concentration was associated with lower birth weight (SD score, adjusted ß: -0.10; 95% CI: -0.15, -0.04; P < 0.001) and placental weight (kilograms, adjusted ß: -10,65; 95% CI: -20.23, -1.07; P = 0.03) and with a higher risk of preeclampsia (adjusted OR: 1.65; 95% CI: 1.10, 2.49; P = 0.02). We observed a 31% decrease in the median plasma t18:1 concentration in our population over time, but the association between the plasma t18:1 concentration standardized per birth year and birth weight was comparable between birth-year cohorts (years 2001-2005).Conclusions: A higher maternal midpregnancy plasma t18:1 concentration was associated with lower birth weight and placental weight and with a higher risk of preeclampsia. Although the intake of TFAs in our population decreased during the inclusion period, the association with adverse pregnancy outcomes was unchanged even at lower maternal plasma t18:1 concentrations.

Associations of maternal folic acid supplementation and folate concentrations during pregnancy with foetal and child head growth: the Generation R Study.

PURPOSE: Folic acid supplementation during pregnancy has been associated with a reduced risk of common neurodevelopmental delays in the offspring. However, it is unclear whether low folate status has effects on the developing brain. We evaluated the associations of maternal folic acid supplementation and folate concentrations during pregnancy with repeatedly measured prenatal and postnatal head circumference in the offspring. METHODS: Within a population-based prospective cohort, we measured maternal plasma folate concentrations at approximately 13 weeks of gestation (90 % range 10.5-17.2) and assessed folic acid supplementation by questionnaire (2001-2005). Up to 11 repeated measures of head circumference were obtained during foetal life (20 and 30 weeks of gestation) and childhood (between birth and age 6 years) in 5866 children (2002-2012). RESULTS: In unadjusted models, foetal head growth was 0.006 SD (95 % CI 0.003; 0.009, P < 0.001) faster per week per 1-SD higher maternal folate concentration. After adjustment for confounders, this association was attenuated to 0.004 SD per week (95 % CI 0.000; 0.007, P = 0.02; estimated absolute difference at birth of 2.7 mm). The association was independent of overall foetal growth. No associations were found between maternal folate concentrations and child postnatal head growth. Preconceptional start of folic acid supplementation was associated with larger prenatal head size, but not with prenatal or postnatal head growth. CONCLUSIONS: Our results suggest an independent, modest association between maternal folate concentrations in early pregnancy and foetal head growth. More research is needed to identify whether specific brain regions are affected and whether effects of folate on foetal head growth influence children's long-term functioning.

Maternal Fatty Acid Status During Pregnancy and Child Autistic Traits: The Generation R Study.

ω-3 and ω-6 polyunsaturated fatty acids are important for brain function and development. We examined whether maternal polyunsaturated fatty acid status during pregnancy affects risk of autistic traits in childhood. Within the Generation R cohort, we measured maternal plasma polyunsaturated fatty acid concentrations and the ω-3:ω-6 ratio in midpregnancy (Rotterdam, the Netherlands, 2001-2005). Child autistic traits at 6 years were assessed by using the Social Responsiveness Scale short form in 4,624 children. A lower maternal ω-3:ω-6 ratio during pregnancy was associated with more autistic traits in the offspring (ß = -0.008, 95% confidence interval: -0.016, -0.001). In particular, a higher total ω-6 and linoleic acid status were associated with more autistic traits (all P's < 0.05). Associations were independent of child intelligence, suggesting that the fatty acid distribution specifically affects the development of autistic traits in addition to general neurodevelopment. Maternal plasma ω-3 status was not associated with child autistic traits and, consistently, neither was prenatal dietary fish intake. Our study shows that a lower prenatal ω-3:ω-6 ratio is associated with more child autistic traits, which is largely accounted for by higher ω-6 instead of lower ω-3 status. These results suggest a biological pathway between maternal fatty acid intake during pregnancy and autistic traits in the offspring.

Maternal fish consumption, fatty acid levels and angiogenic factors: The Generation R Study.

INTRODUCTION: Angiogenic factors, such as placental growth factor (PlGF) and soluble Flt-1 (sFlt-1), are key regulators of placental vascular development. Evidence from in vitro studies indicates that fatty acids can affect angiogenesis. We investigated the associations of maternal fish consumption and fatty acids levels with angiogenic factors during pregnancy, and in cord blood in a large population-based prospective cohort. METHODS: First trimester fish consumption was assessed among 3134 pregnant women using a food-frequency questionnaire. Plasma fatty acid levels were measured in second trimester. Plasma PlGF and sFlt-1 were measured in first and second trimester and in cord blood. Associations of fish consumption or fatty acid levels with angiogenic factors were assessed by multivariable linear regression analyses. RESULTS: There were no consistent associations of total fish or lean fish consumption with levels of PlGF, sFlt-1, or sFlt-1/PlGF ratio. Neither fatty fish nor shellfish were associated with angiogenic factors. Plasma omega-3 polyunsaturated fatty acids, which are the main type of fatty acids in fish, were inconsistently associated with angiogenic factors in second trimester and cord blood. Yet, higher levels of arachidonic acid, an omega-6 polyunsaturated fatty acid, were associated with lower levels of PlGF and sFlt-1. DISCUSSION: We found no consistent associations of fish consumption or fatty acids levels with angiogenic factors in a population with low fish consumption. Studies including populations with higher fish consumption are required to fully grasp the potential effects of maternal fish consumption on placental angiogenesis.

Higher Maternal Plasma n-3 PUFA and Lower n-6 PUFA Concentrations in Pregnancy Are Associated with Lower Childhood Systolic Blood Pressure.

BACKGROUND: Suboptimal maternal diet during pregnancy might lead to fetal cardiovascular adaptations with persistent consequences in the offspring. OBJECTIVE: We assessed the associations of maternal polyunsaturated fatty acid (PUFA) concentrations during pregnancy with childhood blood pressure. METHODS: In a population-based prospective cohort study among 4455 mothers and their children, we measured maternal second-trimester n-3 (ω-3) and n-6 (ω-6) PUFA concentrations in plasma glycerophospholipids and expressed n-3 and n-6 PUFAs as proportions of total PUFAs (wt%). Childhood blood pressure was measured at the median age of 6.0 y (95% range: 5.7-7.9 y). We used linear regression models to assess the associations of maternal PUFA wt% with childhood blood pressure at 6 y. RESULTS: Higher total maternal n-3 PUFA wt% and, specifically, docosahexaenoic acid (DHA; 22:6n-3) wt% were associated with lower childhood systolic blood pressure [differences: -0.28 (95% CI: -0.54, -0.03) and -0.29 mm Hg (95% CI: -0.54, -0.03) per SD increase of total n-3 PUFAs and DHA wt%, respectively], but not with childhood diastolic blood pressure. Total maternal n-6 PUFA wt% was positively associated with childhood systolic blood pressure [differences: 0.36 mm Hg (95% CI: 0.09, 0.62) per SD increase of total n-6 PUFA wt%], but not with childhood diastolic blood pressure. A higher n-6:n-3 PUFA ratio was associated with higher childhood systolic blood pressure (P < 0.05). Pregnancy and childhood characteristics only partly explained the observed associations. CONCLUSIONS: Higher maternal plasma n-3 PUFA and lower n-6 PUFA concentrations during pregnancy are associated with a lower systolic blood pressure in childhood. Further studies are needed to replicate these findings, explore the underlying mechanisms, and examine the long-term cardiovascular consequences.

Folate concentrations during pregnancy and autistic traits in the offspring. The Generation R Study.

In a population-based study, we examined the associations of maternal plasma folate concentrations at 13 weeks of gestation and prenatal folic acid supplement use with autistic traits in the offspring at the age of six years. Parent-reported autistic traits were assessed using the Social Responsiveness Scale short form. Maternal folate was not associated with autistic traits in the offspring. In contrast, prenatal folic acid use was associated with less child autistic traits. Future research should focus on the timing of the potential effect of prenatal folate on the development of autistic traits in combination with clinical diagnosis of autism in the offspring.

Maternal LC-PUFA status during pregnancy and child problem behavior: the Generation R Study.

BACKGROUND: Omega 3 (n-3) and 6 (n-6) long-chain polyunsaturated fatty acids (LC-PUFAs) and the n-3:n-6 ratio are important for brain development. Whether maternal LC-PUFA status during pregnancy affects risk of problem behavior in later childhood is unclear. METHODS: Within a population-based cohort, we measured maternal plasma docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and arachidonic acid (AA) concentrations and n-3:n-6-ratio in mid-pregnancy. Child emotional and behavioral problems at 6 y of age were assessed by parents (child behavior checklist), teachers (teacher report form), and combined parent/teacher report. RESULTS: Higher maternal DHA and n-3:n-6 ratio were associated with fewer child emotional problems using parent (odds ratio (OR)DHA = 0.82; 95% confidence interval (CI): 0.70, 0.96; P = 0.02 and OR(n-3:n-6) = 0.83; 95% CI: 0.71, 0.96; P = 0.01; n = 5,307) and combined parent/teacher scores (ORDHA = 0.79; 95% CI: 0.66, 0.95; P = 0.01 and OR(n-3:n-6) = 0.77; 95% CI: 0.65, 0.92; P < 0.01; n = 2,828). Higher AA was associated with more child behavioral problems using teacher (OR = 1.10; 95% CI: 1.00, 1.20; P = 0.04; n = 3,365) and combined parent/teacher scores (OR = 1.12; 95% CI: 1.02, 1.22; P = 0.02; n = 2,827). Maternal EPA was not associated with child problem behavior. CONCLUSION: Indications of associations of maternal LC-PUFA status with child emotional and behavioral problems were found. Future research is needed to identify LC-PUFA-sensitive periods of fetal brain development by including multiple assessments of prenatal LC-PUFA status.

Maternal urinary iodine concentration in pregnancy and children's cognition: results from a population-based birth cohort in an iodine-sufficient area.

OBJECTIVE: Reports from populations with an insufficient iodine intake suggest that children of mothers with mild iodine deficiency during pregnancy are at risk for cognitive impairments. However, it is unknown whether, even in iodine-sufficient areas, low levels of iodine intake occur that influence cognitive development in the offspring. This study investigated the association between maternal low urinary iodine concentration (UIC) in pregnancy and children's cognition in a population-based sample from a country with an optimal iodine status (the Netherlands). SETTING AND PARTICIPANTS: In 1525 mother-child pairs in a Dutch multiethnic birth cohort, we investigated the relation between maternal UIC<150 µg/g creatinine, assessed <18 weeks gestation and children's cognition. OUTCOMES MEASURES: Non-verbal IQ and language comprehension were assessed during a visit to the research centre using Dutch test batteries when the children were 6 years. RESULTS: In total, 188 (12.3%) pregnant women had UIC<150 µg/g creatinine, with a median UIC equal to 119.3 µg/g creatinine. The median UIC in the group with UIC>150 µg/g creatinine was 322.9 µg/g and in the whole sample 296.5 µg/g creatinine. There was a univariate association between maternal low UIC and children's suboptimum non-verbal IQ (unadjusted OR=1.44, 95% CI 1.02 to 2.02). However, after adjustment for confounders, maternal low UIC was not associated with children's non-verbal IQ (adjusted OR=1.33, 95% CI 0.92 to 1.93). There was no relation between maternal UIC in early pregnancy and children's language comprehension at 6 years. CONCLUSIONS: The lack of a clear association between maternal low UIC and children's cognition probably reflects that low levels of iodine were not frequent and severe enough to affect neurodevelopment. This may result from the Dutch iodine fortification policy, which allows iodised salt to be added to almost all processed food and emphasises the monitoring of iodine intake in the population.

Maternal dietary patterns during pregnancy and child internalising and externalising problems. The Generation R Study.

BACKGROUND & AIMS: Maternal nutritional factors during pregnancy have been linked to foetal brain development and subsequent offspring behaviour. Less is known about associations between maternal dietary patterns and offspring behaviour. METHODS: Within a population-based cohort, we assessed maternal diet using a food frequency questionnaire. Three dietary patterns were derived by means of Principal Component Analysis. Child internalising (emotionally reactive, anxious/depressed or withdrawn, having somatic complaints) and externalising problems (inattention, aggression) were assessed with the Child Behaviour Checklist at 1.5, 3 and 6 years in 3104 children. We assessed the association of maternal Mediterranean, Traditionally Dutch and Confectionary dietary pattern during pregnancy with child internalising and externalising problems. RESULTS: After adjustment, the Mediterranean diet was negatively associated (ORper SD in Mediterranean score = 0.90, 95% CI: 0.83-0.97) and the Traditionally Dutch diet was positively associated with child externalising problems (ORper SD in Traditionally Dutch score = 1.11, 95% CI: 1.03-1.21). Neither diet was associated with internalising problems. CONCLUSIONS: Both low adherence to the Mediterranean diet and high adherence to the Traditionally Dutch diet during pregnancy are associated with an increased risk of child externalising problems. Further research is needed to unravel the effects of nutrient interplay during and after pregnancy on child behavioural development.

Young children's self-reported emotional, behavioral, and peer problems: the Berkeley Puppet Interview.

Adult observers are typically the only informants on emotional and behavioral problems in young children. Although additional information can be provided by child self-report, few validated, structured instruments are available to obtain self-report from young children. The Berkeley Puppet Interview (BPI) has been developed to obtain structured self-reports on multiple domains of mental health and social well-being. This study was the 1st to evaluate the psychometric properties of the BPI in a large sample. We studied 8 a priori scales of the interview in a Dutch community sample of 6,375 children ages 5-7 years. Using confirmatory factor analysis, we demonstrated adequate fit (Tucker-Lewis index = .90; comparative fit index = .90; root-mean-square error of approximation = .03) of a multidimensional model with 50 items loading on 8 latent factors (Depression, Separation Anxiety, Overanxious, Oppositional Defiant, Overt Hostility, Conduct Problems, Bullied by Peers, and Peer Acceptance/Rejection). This model was invariant across gender. Children reported anxiety-related problems more frequently than depressive problems, behavioral problems, or difficulties in peer relations. Reliability analyses showed that 3 broadband scales designated as Internalizing, Externalizing, and Peer Relations were homogeneous constructs (αs = .68-.79). Higher scores on most BPI scales were associated with lower maternal education, lower family income, and non-Western ethnicity. Boys reported more behavioral and peer relation problems, whereas girls reported more emotional problems. The findings indicate that young children from socioeconomically and demographically diverse backgrounds are capable of providing valid, multidimensional information on their emotional, behavioral, and peer relation problems using the BPI. Young children's self-report is a promising addition to existing assessment tools.

Maternal folate status in early pregnancy and child emotional and behavioral problems: the Generation R Study.

BACKGROUND: Maternal prenatal folate status has been linked to neurodevelopmental disorders, but the association with child emotional and behavioral problems is unclear. OBJECTIVES: We assessed the association of maternal folate status during pregnancy with child emotional and behavioral problems. Also, we examined whether any association between folate status and child problems is a consequence of maternal folic acid supplement use or variation in maternal MTHFR genotype. DESIGN: Within a population-based cohort, we measured maternal plasma folate concentrations in early pregnancy and assessed folic acid supplement use by questionnaire. Mothers of European descent were genotyped for the MTHFR 677 C→T polymorphism. Child emotional and behavioral problems were assessed with the Child Behavior Checklist at 3 y in 3209 children. RESULTS: Children of mothers with prenatal folate deficiency were at higher risk of emotional problems (OR: 1.57; 95% CI: 1.03, 2.38) but not behavioral problems (OR: 1.00; 95% CI: 0.64, 1.56) after adjustment for confounders. A higher risk of emotional problems was also found in children whose mothers started using folic acid supplements late or did not use supplements at all (OR: 1.45; 95% CI: 1.14, 1.84) than in children whose mothers started periconceptionally. However, low plasma folate concentrations only partly explained this association (OR: 1.38; 95% CI: 1.08, 1.78). Although related to plasma folate concentrations, maternal MTHFR genotype did not explain the association of folate status with offspring emotional problems. CONCLUSION: Low maternal folate status during early pregnancy is associated with a higher risk of emotional problems in the offspring.