Morphologic development of the first-trimester utero-placental vasculature is positively associated with embryonic and fetal growth: the Rotterdam Periconception Cohort.

STUDY QUESTION: Is morphologic development of the first-trimester utero-placental vasculature associated with embryonic growth and development, fetal growth, and birth weight percentiles? SUMMARY ANSWER: Using the utero-placental vascular skeleton (uPVS) as a new imaging marker, this study reveals morphologic development of the first-trimester utero-placental vasculature is positively associated with embryonic growth and development, fetal growth, and birth weight percentiles. WHAT IS KNOWN ALREADY: First-trimester development of the utero-placental vasculature is associated with placental function, which subsequently impacts embryonic and fetal ability to reach their full growth potential. The attribution of morphologic variations in the utero-placental vascular development, including the vascular structure and branching density, on prenatal growth remains unknown. STUDY DESIGN, SIZE, DURATION: This study was conducted in the VIRTUAL Placental study, a subcohort of 214 ongoing pregnancies, embedded in the prospective observational Rotterdam Periconception Cohort (Predict study). Women were included before 10 weeks gestational age (GA) at a tertiary referral hospital in The Netherlands between January 2017 and March 2018. PARTICIPANTS/MATERIALS, SETTING, METHODS: We obtained three-dimensional power Doppler volumes of the gestational sac including the embryo and the placenta at 7, 9, and 11 weeks of gestation. Virtual Reality-based segmentation and a recently developed skeletonization algorithm were applied to the power Doppler volumes to generate the uPVS and to measure utero-placental vascular volume (uPVV). Absolute vascular morphology was quantified by assigning a morphologic characteristic to each voxel in the uPVS (i.e. end-, bifurcation-crossing-, or vessel point). Additionally, total vascular length (mm) was calculated. The ratios of the uPVS characteristics to the uPVV were calculated to determine the density of vascular branching. Embryonic growth was estimated by crown-rump length and embryonic volume. Embryonic development was estimated by Carnegie stages. Fetal growth was measured by estimated fetal weight in the second and third trimester and birth weight percentiles. Linear mixed models were used to estimate trajectories of longitudinal measurements. Linear regression analysis with adjustments for confounders was used to evaluate associations between trajectories of the uPVS and prenatal growth. Groups were stratified for conception method (natural/IVF-ICSI conceptions), fetal sex (male/female), and the occurrence of placenta-related complications (yes/no). MAIN RESULTS AND THE ROLE OF CHANCE: Increased absolute vascular morphologic development, estimated by positive random intercepts of the uPVS characteristics, is associated with increased embryonic growth, reflected by crown-rump length (endpoints ß = 0.017, 95% CI [0.009; 0.025], bifurcation points ß = 0.012, 95% CI [0.006; 0.018], crossing points ß = 0.017, 95% CI [0.008; 0.025], vessel points ß = 0.01, 95% CI [0.002; 0.008], and total vascular length ß = 0.007, 95% CI [0.003; 0.010], and similarly with embryonic volume and Carnegie stage, all P-values ≤ 0.01. Density of vascular branching was negatively associated with estimated fetal weight in the third trimester (endpoints: uPVV ß = -94.972, 95% CI [-185.245; -3.698], bifurcation points: uPVV ß = -192.601 95% CI [-360.532; -24.670]) and birth weight percentiles (endpoints: uPVV ß = -20.727, 95% CI [-32.771; -8.683], bifurcation points: uPVV ß -51.097 95% CI [-72.257; -29.937], and crossing points: uPVV ß = -48.604 95% CI [-74.246; -22.961])), all P-values < 0.05. After stratification, the associations were observed in natural conceptions specifically. LIMITATION, REASONS FOR CAUTION: Although the results of this prospective observational study clearly demonstrate associations between first-trimester utero-placental vascular morphologic development and prenatal growth, further research is required before we can draw firm conclusions about a causal relationship. WIDER IMPLICATIONS OF THE FINDINGS: Our findings support the hypothesis that morphologic variations in utero-placental vascular development play a role in the vascular mechanisms involved in embryonic and fetal growth and development. Application of the uPVS could benefit our understanding of the pathophysiology underlying placenta-related complications. Future research should focus on the clinical applicability of the uPVS as an imaging marker for the early detection of fetal growth restriction. STUDY FUNDING/COMPETING INTEREST(S): This research was funded by the Department of Obstetrics and Gynecology of the Erasmus MC, University Medical Centre, Rotterdam, The Netherlands. There are no conflicts of interest. TRIAL REGISTRATION NUMBER: Registered at the Dutch Trial Register (NTR6854).

Embryonic morphological development is delayed in pregnancies ending in a spontaneous miscarriage.

STUDY QUESTION: Is there a difference in embryonic morphological development between ongoing pregnancies and live pregnancies ending in a miscarriage? SUMMARY ANSWER: Embryonic morphological development, assessed by the Carnegie stages, is delayed in live pregnancies ending in a miscarriage compared to ongoing pregnancies. WHAT IS KNOWN ALREADY: Pregnancies ending in a miscarriage tend to have smaller embryos and slower heart rates. STUDY DESIGN, SIZE, DURATION: Between 2010 and 2018, 644 women with singleton pregnancies, in the periconception period, were enrolled in a prospective cohort study with follow up until 1 year after delivery. A miscarriage was registered as a non-viable pregnancy before 22 weeks gestational age, defined by an absent heartbeat by ultrasound for a previously reported live pregnancy. PARTICIPANTS/MATERIALS, SETTING, METHODS: Pregnant women with live singleton pregnancies were included and serial three-dimensional transvaginal ultrasound scans were performed. Embryonic morphological development was assessed by the Carnegie developmental stages and evaluated using virtual reality techniques. The embryonic morphology was compared to clinically used growth parameters (i.e. crown-rump length (CRL) and embryonic volume (EV)). Linear mixed models were used to evaluate the association between miscarriage and the Carnegie stages. Logistic regression with generalized estimating equations was used to calculate the odds of a miscarriage after a delay in Carnegie stages. Adjustments were made for potential confounders or covariates and include age, parity, and smoking status. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 611 ongoing pregnancies and 33 pregnancies ending in a miscarriage were included between 7 + 0 and 10 + 3 weeks gestational age, resulting in 1127 assigned Carnegie stages for evaluation. Compared to an ongoing pregnancy, a pregnancy ending in a miscarriage is associated with a lower Carnegie stage (ßCarnegie = -0.824, 95% CI -1.190; -0.458, P < 0.001). A live embryo of a pregnancy ending in a miscarriage will reach the final Carnegie stage with a delay of 4.0 days compared to an ongoing pregnancy. A pregnancy ending in a miscarriage is associated with a smaller CRL (ßCRL = -0.120, 95% CI -0.240; -0.001, P = 0.049) and EV (ßEV = -0.060, 95% CI -0.112; -0.007, P = 0.027). The delay in Carnegie stage increases the odds of a miscarriage by 1.5% per delayed Carnegie stage (ORCarnegie = 1.015, 95% CI 1.002; 1.028, P = 0.028). LIMITATIONS, REASONS FOR CAUTION: We included a relatively small number of pregnancies ending in a miscarriage from a study population that is recruited from a tertiary referral centre. Furthermore, results of genetic testing on the products of the miscarriages or information on the karyotype of the parents were not available. WIDER IMPLICATIONS OF THE FINDINGS: Embryonic morphological development, assessed by the Carnegie stages, is delayed in live pregnancies ending in a miscarriage. In the future, embryonic morphology may be used to estimate the likelihood of a pregnancy continuing to the delivery of a healthy baby. This is of crucial importance for all women but in particular for those at risk of a recurrent pregnancy loss. As part of supportive care, both women and their partners may benefit from information on the prospective outcome of the pregnancy and the timely identification of a miscarriage. STUDY FUNDING/COMPETING INTEREST(S): The work was funded by the Department of Obstetrics and Gynaecology, Erasmus MC, University Medical Centre, Rotterdam, The Netherlands. The authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

First trimester anomaly scan using virtual reality (VR FETUS study): study protocol for a randomized clinical trial.

BACKGROUND: In recent years it has become clear that fetal anomalies can already be detected at the end of the first trimester of pregnancy by two-dimensional (2D) ultrasound. This is why increasingly in developed countries the first trimester anomaly scan is being offered as part of standard care. We have developed a Virtual Reality (VR) approach to improve the diagnostic abilities of 2D ultrasound. Three-dimensional (3D) ultrasound datasets are used in VR assessment, enabling real depth perception and unique interaction. The aim of this study is to investigate whether first trimester 3D VR ultrasound is of additional value in terms of diagnostic accuracy for the detection of fetal anomalies. Health-related quality of life, cost-effectiveness and also the perspective of both patient and ultrasonographer on the 3D VR modality will be studied. METHODS: Women in the first trimester of a high risk pregnancy for a fetus with a congenital anomaly are eligible for inclusion. This is a randomized controlled trial with two intervention arms. The control group receives 'care as usual': a second trimester 2D advanced ultrasound examination. The intervention group will undergo an additional first trimester 2D and 3D VR ultrasound examination. Following each examination participants will fill in validated questionnaires evaluating their quality of life and healthcare related expenses. Participants' and ultrasonographers' perspectives on the 3D VR ultrasound will be surveyed. The primary outcome will be the detection of fetal anomalies. The additional first trimester 3D VR ultrasound examination will be compared to 'care as usual'. Neonatal or histopathological examinations are considered the gold standard for the detection of congenital anomalies. To reach statistical significance and 80% power with a detection rate of 65% for second trimester ultrasound examination and 70% for the combined detection of first trimester 3D VR and second trimester ultrasound examination, a sample size of 2800 participants is needed. DISCUSSION: First trimester 3D VR detection of fetal anomalies may improve patients' quality of life through reassurance or earlier identification of malformations. Results of this study will provide policymakers and healthcare professionals with the highest level of evidence for cost-effectiveness of first trimester ultrasound using a 3D VR approach. TRIAL REGISTRATION: Dutch Trial Registration number NTR6309 , date of registration 26 January 2017.

CCL22 is a biomarker of cartilage injury and plays a functional role in chondrocyte apoptosis.

BACKGROUND: Osteoarthritis (OA) is one of the leading causes of disability worldwide. Previous history of knee injury is a significant risk factor for OA. It has been established that low-level chronic inflammation plays a pivotal role in the onset and pathogenesis of OA. The primary aim of this research was to determine if a history of knee joint injury is associated with systemic inflammation. A secondary aim was to determine if systemic inflammation is related to knee pain and joint structure. METHODS: Differences in serum cytokine association networks, knee joint structural changes (MRI), and self-reported pain (i.e., Knee Injury and Osteoarthritis Outcome Score Pain subscale, KOOSPAIN and Intermittent and Constant Osteoarthritis Pain score, ICOAP) between individuals who had sustained a youth (aged 15-26 years) sport-related knee injury 3-10 years previously and age- and sex-matched controls were examined. Proteins of interest were also examined in an OA rat model. RESULTS: Cytokine association networks were found to differ significantly between study groups, yet no significant associations were found between networks and KOOSPAIN or MRI-defined OA. A group of cytokines (MCP1/CCL2, CCL22 and TNFα) were differentially associated with other cytokines between study groups. In a pre-clinical rat OA model, serum CCL22 levels were associated with pain (r = 0.255, p = 0.045) and structural changes to the cartilage. CCL22 expression was also observed in human OA cartilage and furthermore, CCL22 induced apoptosis of isolated human chondrocytes. DISCUSSION: These results suggest that CCL22 may be an early factor in the onset/pathogenic process of cartilage degeneration and/or related to pain OA.

Wright-Fisher exact solver (WFES): scalable analysis of population genetic models without simulation or diffusion theory.

Motivation: The simplifying assumptions that are used widely in theoretical population genetics may not always be appropriate for empirical population genetics. General computational approaches that do not require the assumptions of classical theory are therefore quite desirable. One such general approach is provided by the theory of absorbing Markov chains, which can be used to obtain exact results by directly analyzing population genetic Markov models, such as the classic bi-allelic Wright-Fisher model. Although these approaches are sometimes used, they are usually forgone in favor of simulation methods, due to the perception that they are too computationally burdensome. Here we show that, surprisingly, direct analysis of virtually any Markov chain model in population genetics can be made quite efficient by exploiting transition matrix sparsity and by solving restricted systems of linear equations, allowing a wide variety of exact calculations (within machine precision) to be easily and rapidly made on modern workstation computers. Results: We introduce Wright-Fisher Exact Solver (WFES), a fast and scalable method for direct analysis of Markov chain models in population genetics. WFES can rapidly solve for both long-term and transient behaviours including fixation and extinction probabilities, expected times to fixation or extinction, sojourn times, expected allele age and variance, and others. Our implementation requires only seconds to minutes of runtime on modern workstations and scales to biological population sizes ranging from humans to model organisms. Availability and Implementation: The code is available at https://github.com/dekoning-lab/wfes. Contact: jason.dekoning@ucalgary.ca. Supplementary information: Supplementary data are available at Bioinformatics online.

EzMap: a simple pipeline for reproducible analysis of the human virome.

SUMMARY: In solid-organ transplant recipients, a delicate balance between immunosuppression and immunocompetence must be achieved, which can be difficult to monitor in real-time. Shotgun sequencing of cell-free DNA (cfDNA) has been recently proposed as a new way to indirectly assess immune function in transplant recipients through analysis of the status of the human virome. To facilitate exploration of the utility of the human virome as an indicator of immune status, and to enable rapid, straightforward analyses by clinicians, we developed a fully automated computational pipeline, EzMap, for performing metagenomic analysis of the human virome. EzMap combines a number of tools to clean, filter, and subtract WGS reads by mapping to a reference human assembly. The relative abundance of each virus present is estimated using a maximum likelihood approach that accounts for genome size, and results are presented with interactive visualizations and taxonomy-based summaries that enable rapid insights. The pipeline is automated to run on both workstations and computing clusters for all steps. EzMap automates an otherwise tedious and time-consuming protocol and aims to facilitate rapid and reproducible insights from cfDNA. AVAILABILITY AND IMPLEMENTATION: EzMap is freely available at https://github.com/dekoning-lab/ezmap. CONTACT: jason.dekoning@ucalgary.ca. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Breast and Tumour Volume Measurements in Breast Cancer Patients Using 3-D Automated Breast Volume Scanner Images.

BACKGROUND: The resection volume in relation to the breast volume is known to influence cosmetic outcome following breast-conserving therapy. It was hypothesised that three-dimensional ultrasonography (3-D US) could be used to preoperatively assess breast and tumour volume and show high association with histopathological measurements. METHODS: Breast volume by the 3D-US was compared to the water displacement method (WDM), mastectomy specimen weight, 3-D MRI and three different calculations for breast volume on mammography. Tumour volume by the 3-D US was compared to the histopathological tumour volume and 3-D MRI. Relatedness was based on the intraclass correlation coefficient (ICC) with corresponding 95% confidence interval (95% CI). Bland-Altman plots were used to graphically display the agreement for the different assessment techniques. All measurements were performed by one observer. RESULTS: A total of 36 patients were included, 20 and 23 for the evaluation of breast and tumour volume (ductal invasive carcinomas), respectively. 3-D US breast volume showed 'excellent' association with WDM, ICC 0.92 [95% CI (0.80-0.97)]. 3-D US tumour volume showed a 'excellent' association with histopathological tumour volume, ICC 0.78 [95% CI (0.55-0.91)]. Bland-Altman plots showed an increased overestimation in lager tumour volumes measured by 3-D MRI compared to histopathological volume. CONCLUSIONS: 3-D US showed a high association with gold standard WDM for the preoperative assessment of breast volume and the histopathological measurement of tumour volume. 3-D US is an patient-friendly preoperative available technique to calculate both breast volume and tumour volume. Volume measurements are promising in outcome prediction of intended breast-conserving treatment.

Periconceptional maternal one-carbon biomarkers are associated with embryonic development according to the Carnegie stages.

STUDY QUESTION: Is periconceptional maternal one-carbon (I-C) metabolism associated with embryonic morphological development in non-malformed ongoing pregnancies? SUMMARY ANSWER: Serum vitamin B12, red blood cell (RBC) folate and plasma total homocysteine (tHcy) are associated with embryonic development according to the Carnegie stages. WHAT IS KNOWN ALREADY: Derangements in maternal I-C metabolism affect reproductive and pregnancy outcomes, as well as future health of the offspring. STUDY DESIGN, SIZE, DURATION: Between 2010 and 2014, women with singleton ongoing pregnancies were enrolled in a prospective periconceptional cohort study. PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 234 pregnancies, including 138 spontaneous or IUI pregnancies with strict pregnancy dating and 96 pregnancies derived from IVF, ICSI or cryopreserved embryo transfer (IVF/ICSI pregnancies), underwent longitudinal transvaginal three-dimensional ultrasound (3D US) scans from 6+0 up to 10+2 weeks of gestation. Carnegie stages were defined using internal and external morphologic criteria in a virtual reality system. Maternal venous blood samples were collected at enrollment for serum vitamin B12, RBC folate and plasma tHcy assessment. Associations between biomarker concentrations and longitudinal Carnegie stages were investigated using linear mixed models. MAIN RESULTS AND THE ROLE OF CHANCE: We performed a median of three 3D US scans per pregnancy (range 1-5) resulting in 600 good quality data sets for the Carnegie stage annotation (80.5%). Vitamin B12 was positively associated with embryonic development in the total study population (ß = 0.001 (95% CI: 0.000; 0.002), P < 0.05) and in the subgroup of strictly dated spontaneous pregnancies (ß = 0.002 (95% CI: 0.001; 0.003), P < 0.05). Low vitamin B12 concentrations (-2SD, 73.4 pmol/l) were associated with delayed embryonic development by 1.4 days (95% CI: 1.3-1.4) compared with high concentrations (+2SD, 563.1 pmol/l). RBC folate was positively associated with Carnegie stages only in IVF/ICSI pregnancies (ß = 0.001 (95% CI: 0.0005; 0.0015), P < 0.05). In this group, low RBC folate concentrations (-2SD, 875.4 nmol/l) were associated with a 1.8-day delay (95% CI: 1.7-1.8) in development compared with high concentrations (+2SD, 2119.9 nmol/l). tHcy was negatively associated with embryonic development in the total study population (ß = -0.08 (95% CI: -0.14; -0.02), P < 0.01), as well as in the IVF/ICSI subgroup (ß = -0.08 (95% CI: -0.15; -0.01), P < 0.05). High tHcy concentrations (+2SD, 10.4 µmol/l) were associated with a delay of 1.6 days (95% CI: 1.5-1.7) in embryonic development compared with low concentrations (-2SD, 3.0 µmol/l). LIMITATIONS, REASONS FOR CAUTION: The study was performed in a tertiary care center, resulting in high rates of folic acid supplement use and comorbidity that may reduce the external validity of our findings. WIDER IMPLICATIONS OF THE FINDINGS: In periconceptional care, maternal I-C biomarkers should be taken into account as predictors of embryonic morphological development. Combining embryonic size measurements with morphological assessment could better define normal embryonic development. STUDY FUNDING/COMPETING INTEREST(S): The work was funded by the Department of Obstetrics and Gynaecology, Erasmus MC, University Medical Centre, Rotterdam, The Netherlands. RPMST is CSO of the startup company Slimmere Zorg and CEO of eHealth Care Solutions. The authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER: Not applicable.

TUmor-volume to breast-volume RAtio for improving COSmetic results in breast cancer patients (TURACOS); a randomized controlled trial.

BACKGROUND: Cosmetic result following breast conserving surgery (BCS) for cancer influences quality of life and psychosocial functioning in breast cancer patients. A preoperative prediction of expected cosmetic result following BCS is not (yet) standard clinical practice and therefore the choice for either mastectomy or BCS is still subjective. Recently, we showed that tumour volume to breast volume ratio as well as tumour location in the breast are independent predictors of superior cosmetic result following BCS. Implementation of a prediction model including both factors, has not been studied in a prospective manner. This study aims to improve cosmetic outcome by implementation of a prediction model in the treatment decision making for breast cancer patients opting for BCS. METHODS/DESIGN: Multicentre, single-blinded, randomized controlled trial comparing standard preoperative work-up to a preoperative work-up with addition of the prediction model. Tumour volume to breast volume ratio and tumour location in the breast will be used to predict cosmetic outcome in invasive breast cancer patients opting for BCS. Three dimensional (3D)-ultrasonography will be used to measure the tumour volume to breast volume ratio needed for the prediction model. Sample size was estimated based on a 14% improvement in incidence of superior cosmetic result one year after BCS (71% in the control group versus 85% in the intervention group). Primarily cosmetic outcome will be evaluated by a 6-member independent panel. Secondary endpoints include; (1) patient reported outcome measured by BREAST-Q, EORTC-QLQ-C30/BR23 and EQ-5D-5 L (2) cosmetic outcome as assessed through the BCCT.core software, (3) radiation-induced reaction (4) surgical treatment performed, (5) pathological result and (6) cost-effectiveness. Follow-up data will be collected for 3 years after surgery or finishing radiotherapy. DISCUSSION: This randomized controlled trial examines the value of a preoperative prediction model for the treatment-decision making. It aims for a superior cosmetic result in breast cancer patients opting for BCS. We expect improvement of patients' quality of life and psychosocial functioning in a cost-effective way. TRIAL REGISTRATION: Prospectively registered, February 17th 2015, at 'Nederlands Trialregister - NTR4997 '.

Periconceptional maternal 'high fish and olive oil, low meat' dietary pattern is associated with increased embryonic growth: The Rotterdam Periconceptional Cohort (Predict) Study.

OBJECTIVE: To investigate the association between periconceptional maternal dietary pattern and first-trimester embryonic growth. METHODS: This was a prospective cohort study of 228 women with a singleton ongoing pregnancy, of which 135 were strictly dated spontaneous pregnancies and 93 were pregnancies achieved after in-vitro fertilization or intracytoplasmatic sperm injection (IVF/ICSI). All women underwent serial transvaginal three-dimensional ultrasound (3D-US) examinations from 6 + 0 to 13 + 0 weeks' gestation. Crown-rump length (CRL) and embryonic volume (EV) measurements were performed using a virtual reality system. Information on periconceptional maternal dietary intake was collected via food frequency questionnaires. Principal component analysis was performed to identify dietary patterns. Associations between dietary patterns and CRL and EV trajectories were investigated using linear mixed models adjusted for potential confounders. RESULTS: A median of five (range, one to seven) 3D-US scans per pregnancy were performed. Of 1162 datasets, quality was sufficient to perform CRL measurements in 991 (85.3%) and EV measurements in 899 (77.4%). A dietary pattern comprising high intake of fish and olive oil and a very low intake of meat was identified as beneficial for embryonic growth. In strictly dated spontaneous pregnancies, strong adherence to the 'high fish and olive oil, low meat' dietary pattern was associated with a 1.9 mm (95% CI, 0.1-3.63 mm) increase in CRL (+14.6%) at 7 weeks and a 3.4 mm (95% CI, 0.2-7.81 mm) increase (+6.9%) at 11 weeks, whereas EV increased by 0.06 cm3 (95% CI, 0.01-0.13 cm3 ) (+20.4%) at 7 weeks and 1.43 cm3 (95% CI, 0.99-1.87 cm3 ) (+14.4%) at 11 weeks. No significant association was observed in the total study population or in the IVF/ICSI subgroup. CONCLUSION: Periconceptional maternal adherence to a high fish and olive oil, low meat dietary pattern is positively associated with embryonic growth in spontaneously conceived pregnancies. Copyright © 2017 ISUOG. Published by John Wiley & Sons Ltd.

Growth trajectories of the human embryonic head and periconceptional maternal conditions.

STUDY QUESTION: Can growth trajectories of the human embryonic head be created using 3D ultrasound (3D-US) and virtual reality (VR) technology, and be associated with second trimester fetal head size and periconceptional maternal conditions? SUMMARY ANSWER: Serial first trimester head circumference (HC) and head volume (HV) measurements were used to create reliable growth trajectories of the embryonic head, which were significantly associated with fetal head size and periconceptional maternal smoking, age and ITALIC! in vitro fertilization (IVF)/intra-cytoplasmic sperm injection (ICSI) treatment. WHAT IS KNOWN ALREADY: Fetal growth is influenced by periconceptional maternal conditions. STUDY DESIGN, SIZE, DURATION: We selected 149 singleton pregnancies with a live born non-malformed fetus from the Rotterdam periconception cohort. PARTICIPANTS/MATERIALS, SETTING, METHODS: Bi-parietal diameter and occipital frontal diameter to calculate HC, HV and crown-rump length (CRL) were measured weekly between 9 + 0 and 12 + 6 weeks gestational age (GA) using 3D-US and VR. Fetal HC was obtained from second trimester structural anomaly scans. Growth trajectories of the embryonic head were created with general additive models and linear mixed models were used to estimate associations with maternal periconceptional conditions as a function of GA and CRL, respectively. MAIN RESULTS: A total of 303 3D-US images of 149 pregnancies were eligible for embryonic head measurements (intra-class correlation coefficients >0.99). Associations were found between embryonic HC and fetal HC ( ITALIC! ρ = 0.617, ITALIC! P < 0.001) and between embryonic HV and fetal HC ( ITALIC! ρ = 0.660, ITALIC! P < 0.001) in ITALIC! Z-scores. Maternal periconceptional smoking was associated with decreased, and maternal age and IVF/ICSI treatment with increased growth trajectories of the embryonic head measured by HC and HV (All ITALIC! P < 0.05). LIMITATIONS, REASONS FOR CAUTION: The consequences of the small effect sizes for neurodevelopmental outcome need further investigation. As the study population consists largely of tertiary hospital patients, external validity should be studied in the general population. WIDER IMPLICATIONS OF THE FINDINGS: Assessment of growth trajectories of the embryonic head may be of benefit in future early antenatal care. STUDY FUNDING/COMPETING INTERESTS: This study was funded by the Department of Obstetrics and Gynaecology, Erasmus MC University Medical Centre and Sophia Foundation for Medical Research, Rotterdam, The Netherlands (SSWO grant number 644). No competing interests are declared.

The periconception maternal cardiovascular risk profile influences human embryonic growth trajectories in IVF/ICSI pregnancies.

STUDY QUESTION: Is the maternal cardiovascular (CV) risk profile associated with human embryonic growth trajectories and does the mode of conception affect this association? SUMMARY ANSWER: This small study suggests that the maternal CV risk profile is inversely associated with first trimester embryonic growth trajectories in in vitro fertilization (IVF)/intra-cytoplasmic sperm injection (ICSI) pregnancies, but not in spontaneously conceived pregnancies. WHAT IS KNOWN ALREADY: Maternal high-blood pressure and smoking affect placental function, accompanied by increased risk of fetal growth restriction and low-birthweight. Mothers who experience pregnancies complicated by fetal growth restriction are at increased risk of CV disease in later life. STUDY DESIGN, SIZE, DURATION: In a prospective periconception birth cohort conducted in a tertiary hospital, 111 singleton ongoing pregnancies with reliable pregnancy dating, no pre-existing maternal disease and no malformed live borns were investigated. PARTICIPANTS/MATERIALS, SETTINGS, METHODS: Spontaneously conceived pregnancies with a reliable first day of the last menstrual period and a regular menstrual cycle of 25-31 days only (n = 66) and IVF/ICSI pregnancies (n = 45) were included. Women underwent weekly three-dimensional ultrasound scans (3D US) from 6- to 13-week gestational age. To estimate embryonic growth, serial crown-rump length (CRL) measurements were performed using the V-Scope software in a BARCO I-Space. Maternal characteristics and CV risk factors were collected by self-administered questionnaires. The CV risk profile was created based on a score of risk factors, including maternal age, body-mass index, CV disease in the family, diet and smoking. Quartiles of the CV risk score were calculated. Associations between the CV risk score and embryonic growth were assessed using square root transformed CRL in multivariable linear mixed model analyses. MAIN RESULTS AND THE ROLE OF CHANCE: From the 111 included pregnancies, 696 3D US data sets were obtained of which 637 (91.5%) CRLs could be measured. In the total group, The CV risk score was inversely, but not significantly associated with embryonic growth (-0.03√mm; P = 0.291). Stratified by mode of conception, the CV risk score was inversely and significantly associated with embryonic growth (ß = -0.04√mm; P = 0.025, adjusted for possible confounders) in the IVF/ICSI group. Compared with the first quartile, embryos in the upper quartile were 10.4% smaller at 6(+0) weeks (4.4 versus 4.9 mm) and 3.1% smaller at 12(+0) weeks (56.5 versus 58.4 mm) of gestation. Although the CV risk score was slightly, but significantly, higher in women conceiving spontaneously compared with those undergoing IVF/ICSI treatment [CV risk score = 2.06 (SD: 1.23) and 1.60 (SD: 1.15), respectively], no association was established with embryonic growth in that particular group. LIMITATIONS, REASONS FOR CAUTION: Participants included in the present cohort are women with a singleton ongoing pregnancy without any pre-existing disease and selected from a tertiary hospital. Hence, they represent a selected group of women. Larger and population-based periconception birth cohort studies are recommended to demonstrate external validity. WIDER IMPLICATIONS OF THE FINDINGS: Differences in embryonic growth between pregnancies conceived spontaneously and after IVF/ICSI treatment in relation with CV risk factors substantiate the importance of more investigation into differences in sensitivity of endometrial, endothelial, placental and embryonic tissues. STUDY FUNDING/COMPETING INTERESTS: Funded by the Department of Obstetrics and Gynaecology, Erasmus MC, University Medical Centre, Rotterdam, the Netherlands. The authors declare no conflict of interest.

Agreement and reliability of pelvic floor measurements during rest and on maximum Valsalva maneuver using three-dimensional translabial ultrasound and virtual reality imaging.

OBJECTIVES: Imaging of the levator ani hiatus provides valuable information for the diagnosis and follow-up of patients with pelvic organ prolapse (POP). This study compared measurements of levator ani hiatal volume during rest and on maximum Valsalva, obtained using conventional three-dimensional (3D) translabial ultrasound and virtual reality imaging. Our objectives were to establish their agreement and reliability, and their relationship with prolapse symptoms and POP quantification (POP-Q) stage. METHODS: One hundred women with an intact levator ani were selected from our tertiary clinic database. Information on clinical symptoms were obtained using standardized questionnaires. Ultrasound datasets were analyzed using a rendered volume with a slice thickness of 1.5 cm, at the level of minimal hiatal dimensions, during rest and on maximum Valsalva. The levator area (in cm(2) ) was measured and multiplied by 1.5 to obtain the levator ani hiatal volume (in cm(3) ) on conventional 3D ultrasound. Levator ani hiatal volume (in cm(3) ) was measured semi-automatically by virtual reality imaging using a segmentation algorithm. Twenty patients were chosen randomly to analyze intra- and interobserver agreement. RESULTS: The mean difference between levator hiatal volume measurements on 3D ultrasound and by virtual reality was 1.52 cm(3) (95% CI, 1.00-2.04 cm(3) ) at rest and 1.16 cm(3) (95% CI, 0.56-1.76 cm(3) ) during maximum Valsalva (P < 0.001). Both intra- and interobserver intraclass correlation coefficients were ≥ 0.96 for conventional 3D ultrasound and > 0.99 for virtual reality. Patients with prolapse symptoms or POP-Q Stage ≥ 2 had significantly larger hiatal measurements than those without symptoms or POP-Q Stage < 2. CONCLUSIONS: Levator ani hiatal volume at rest and on maximum Valsalva is significantly smaller when using virtual reality compared with conventional 3D ultrasound; however, this difference does not seem clinically important. Copyright © 2015 ISUOG. Published by John Wiley & Sons Ltd.

The king cobra genome reveals dynamic gene evolution and adaptation in the snake venom system.

Snakes are limbless predators, and many species use venom to help overpower relatively large, agile prey. Snake venoms are complex protein mixtures encoded by several multilocus gene families that function synergistically to cause incapacitation. To examine venom evolution, we sequenced and interrogated the genome of a venomous snake, the king cobra (Ophiophagus hannah), and compared it, together with our unique transcriptome, microRNA, and proteome datasets from this species, with data from other vertebrates. In contrast to the platypus, the only other venomous vertebrate with a sequenced genome, we find that snake toxin genes evolve through several distinct co-option mechanisms and exhibit surprisingly variable levels of gene duplication and directional selection that correlate with their functional importance in prey capture. The enigmatic accessory venom gland shows a very different pattern of toxin gene expression from the main venom gland and seems to have recruited toxin-like lectin genes repeatedly for new nontoxic functions. In addition, tissue-specific microRNA analyses suggested the co-option of core genetic regulatory components of the venom secretory system from a pancreatic origin. Although the king cobra is limbless, we recovered coding sequences for all Hox genes involved in amniote limb development, with the exception of Hoxd12. Our results provide a unique view of the origin and evolution of snake venom and reveal multiple genome-level adaptive responses to natural selection in this complex biological weapon system. More generally, they provide insight into mechanisms of protein evolution under strong selection.

The Burmese python genome reveals the molecular basis for extreme adaptation in snakes.

Snakes possess many extreme morphological and physiological adaptations. Identification of the molecular basis of these traits can provide novel understanding for vertebrate biology and medicine. Here, we study snake biology using the genome sequence of the Burmese python (Python molurus bivittatus), a model of extreme physiological and metabolic adaptation. We compare the python and king cobra genomes along with genomic samples from other snakes and perform transcriptome analysis to gain insights into the extreme phenotypes of the python. We discovered rapid and massive transcriptional responses in multiple organ systems that occur on feeding and coordinate major changes in organ size and function. Intriguingly, the homologs of these genes in humans are associated with metabolism, development, and pathology. We also found that many snake metabolic genes have undergone positive selection, which together with the rapid evolution of mitochondrial proteins, provides evidence for extensive adaptive redesign of snake metabolic pathways. Additional evidence for molecular adaptation and gene family expansions and contractions is associated with major physiological and phenotypic adaptations in snakes; genes involved are related to cell cycle, development, lungs, eyes, heart, intestine, and skeletal structure, including GRB2-associated binding protein 1, SSH, WNT16, and bone morphogenetic protein 7. Finally, changes in repetitive DNA content, guanine-cytosine isochore structure, and nucleotide substitution rates indicate major shifts in the structure and evolution of snake genomes compared with other amniotes. Phenotypic and physiological novelty in snakes seems to be driven by system-wide coordination of protein adaptation, gene expression, and changes in the structure of the genome.

First trimester trophoblast and placental bed vascular volume measurements in IVF or IVF/ICSI pregnancies.

STUDY QUESTION: Are first trimester trophoblast volume (TV) and placental bed vascular volume (PBVV) different in IVF or IVF/ICSI pregnancies in comparison with spontaneously conceived pregnancies? SUMMARY ANSWER: Any possible abnormal placentation in IVF or IVF/ICSI pregnancies in comparison with spontaneously conceived pregnancies is not detected by a difference in PBVV or TV at an early gestational age (GA). WHAT IS KNOWN ALREADY: Assisted reproductive technology pregnancies have been associated with an increased risk of placenta-related adverse pregnancy outcomes. It is unclear whether these effects originate from infertility or from the technique itself. STUDY DESIGN, SIZE, DURATION: We performed a retrospective cohort study in which 154 pregnant patients qualified for participation. PARTICIPANTS/MATERIALS, SETTING, METHODS: Out of 154 pregnant patients, 84 conceived spontaneously and 70 conceived after IVF or IVF/ICSI. We determined the TV at 10 weeks GA by Virtual Organ Computer-aided AnaLysis measuring application and the PBVV at 12 weeks GA by the virtual reality operating system of BARCO I-Space in both subgroups. The investigators were blinded to the mode of conception during the measurements. Analysis was limited to singleton pregnancies with only one sac ever detectable. MAIN RESULTS AND THE ROLE OF CHANCE: There were no differences in TV (mean 42.7, SD 15.9 versus mean 41.2, SD 13.9, P = 0.70) and PBVV (mean 27.6, SD 16.9 versus mean 24.8, SD 19.9, P = 0.20) between IVF or IVF/ICSI pregnancies and spontaneously conceived pregnancies. There was a significant correlation between TV and PBVV (rs = 0.283, P = 0.004). LIMITATIONS, REASONS FOR CAUTION: The limitations of the present study concern the small size of the study groups. WIDER IMPLICATIONS OF THE FINDINGS: IVF or IVF/ICSI does not seem to be associated with abnormal placentation. STUDY FUNDING/COMPETING INTERESTS: This study was financially supported by the Erasmus Trustfonds, the Meindert de Hoop foundation and the Fonds NutsOhra. No competing interests are declared.

The influence of IVF/ICSI treatment on human embryonic growth trajectories.

STUDY QUESTION: Is in vitro fertilization treatment with or without intracytoplasmatic sperm injection (IVF/ICSI) associated with changes in first and second trimester embryonic and fetal growth trajectories and birthweight in singleton pregnancies? SUMMARY ANSWER: Embryonic and fetal growth trajectories and birthweight are not significantly different between pregnancies conceived with IVF/ICSI treatment and spontaneously conceived pregnancies with reliable pregnancy dating. WHAT IS KNOWN ALREADY: IVF/ICSI treatment has been associated with increased risks of preterm birth, fetal growth restriction and low birthweight. Decreased first-trimester crown-rump length (CRL) in the general population has been inversely associated with the same adverse pregnancy outcomes. STUDY DESIGN, SIZE, DURATION: In a prospective periconception birth cohort study conducted in a tertiary centre, 146 singleton pregnancies with reliable pregnancy dating and nonmalformed live borns were investigated, comprised of 88 spontaneous and 58 IVF/ICSI pregnancies. PARTICIPANTS/MATERIALS, SETTING, METHODS: Serial 3D ultrasound scans were performed from 6 to 12 weeks of gestation. As estimates of embryonic growth, CRL and embryonic volume (EV) were measured using the I-Space virtual reality system. General characteristics were obtained from self-administered questionnaires at enrolment. Fetal growth parameters at 20 weeks and birthweight were obtained from medical records. To assess associations between IVF/ICSI and embryonic growth trajectories, estimated fetal weight and birthweight, stepwise linear mixed model analyses and linear regression analyses were performed using square root transformed CRL and fourth root transformed EV. MAIN RESULTS AND THE ROLE OF CHANCE: In 146 pregnancies, 934 ultrasound scans were performed of which 849 (90.9%) CRLs and 549 (58.8%) EVs could be measured. Embryonic growth trajectories were comparable between IVF/ICSI pregnancies and spontaneously conceived pregnancies (CRL: ßIVF/ICSI = 0.10√mm; P = 0.10; EV: ßIVF/ICSI = 0.03(4)√cm³; P = 0.13). Estimated fetal weight and birthweight were also comparable between both groups (ßIVF/ICSI = 6 g; P = 0.36 and ßIVF/ICSI = 80 g; P = 0.24, respectively). LIMITATIONS, REASONS FOR CAUTION: Variations in embryonic growth trajectories of spontaneously conceived pregnancies with reliable pregnancy dating may partially be a result of less precise pregnancy dating and differences in endometrium receptivity compared with IVF/ICSI pregnancies. WIDER IMPLICATIONS OF THE FINDINGS: The absence of a significant difference in embryonic and fetal growth trajectories suggests safety of IVF/ICSI treatment with regard to early embryonic growth. However, further research is warranted to ascertain the influence of IVF/ICSI treatments in a larger study population, and to estimate the impact of the underlying causes of the subfertility and other periconceptional exposures on human embryonic and fetal growth trajectories. FUNDING STATEMENT: This study was supported by the Department of Obstetrics and Gynaecology of the Erasmus MC, University Medical Centre. CONFLICT OF INTEREST: No competing interests are declared.

An optimal periconception maternal folate status for embryonic size: the Rotterdam Predict study.

OBJECTIVE: To investigate the association between periconception maternal folate status and embryonic size. DESIGN: Prospective periconception cohort study. SETTING: Erasmus University Medical Centre, Rotterdam, the Netherlands. POPULATION: Seventy-seven singleton pregnancies recruited in 2009 and 2010. METHODS: We recruited women before 8 weeks of gestation and performed weekly three-dimensional ultrasound scans from enrolment up to 13 weeks of gestation. As a measure of embryonic growth, crown-rump length (CRL) measurements were performed using V-Scope software in the BARCO I-Space. Maternal blood was collected to determine first-trimester long-term red blood cell (RBC) folate status. Non-malformed live births were included in the analysis. We calculated quartiles of RBC folate, square root-transformed CRL data and performed multivariable linear mixed model analyses. MAIN OUTCOME MEASURES: Serial first-trimester CRL measurements. RESULTS: In total, 484 ultrasound scans were performed in 77 women, in 440 (90.7%) of which CRLs could be measured. RBC folate in the third quartile (1513-1812 nmol/l) was significantly associated with an increased CRL compared with the first two quartiles (814-1512 nmol/l) and the upper quartile (1813-2936 nmol/l; P(overall) = 0.03; adjusted for gestational age, smoking, body mass index and fetal sex). Compared with the third quartile, embryos in the upper quartile were 24.2% smaller at 6(+0) weeks [4.1 mm (95% confidence interval 3.5, 4.7) versus 5.4 mm (95% confidence interval 4.8, 6.1)] and 7.6% smaller at 12(+0) weeks [55.1 mm (95% confidence interval 52.9, 57.3) versus 59.6 mm (95% confidence interval 57.4, 62.0)] of gestation. CONCLUSIONS: This study suggests that a very high maternal periconception folate status is associated with reduced embryonic size. Whether these effects are beneficial or harmful requires further investigation.

Repetitive elements may comprise over two-thirds of the human genome.

Transposable elements (TEs) are conventionally identified in eukaryotic genomes by alignment to consensus element sequences. Using this approach, about half of the human genome has been previously identified as TEs and low-complexity repeats. We recently developed a highly sensitive alternative de novo strategy, P-clouds, that instead searches for clusters of high-abundance oligonucleotides that are related in sequence space (oligo "clouds"). We show here that P-clouds predicts >840 Mbp of additional repetitive sequences in the human genome, thus suggesting that 66%-69% of the human genome is repetitive or repeat-derived. To investigate this remarkable difference, we conducted detailed analyses of the ability of both P-clouds and a commonly used conventional approach, RepeatMasker (RM), to detect different sized fragments of the highly abundant human Alu and MIR SINEs. RM can have surprisingly low sensitivity for even moderately long fragments, in contrast to P-clouds, which has good sensitivity down to small fragment sizes (∼25 bp). Although short fragments have a high intrinsic probability of being false positives, we performed a probabilistic annotation that reflects this fact. We further developed "element-specific" P-clouds (ESPs) to identify novel Alu and MIR SINE elements, and using it we identified ∼100 Mb of previously unannotated human elements. ESP estimates of new MIR sequences are in good agreement with RM-based predictions of the amount that RM missed. These results highlight the need for combined, probabilistic genome annotation approaches and suggest that the human genome consists of substantially more repetitive sequence than previously believed.

Phylogenetics, likelihood, evolution and complexity.

SUMMARY: Phylogenetics, likelihood, evolution and complexity (PLEX) is a flexible and fast Bayesian Markov chain Monte Carlo software program for large-scale analysis of nucleotide and amino acid data using complex evolutionary models in a phylogenetic framework. The program gains large speed improvements over standard approaches by implementing 'partial sampling of substitution histories', a data augmentation approach that can reduce data analysis times from months to minutes on large comparative datasets. A variety of nucleotide and amino acid substitution models are currently implemented, including non-reversible and site-heterogeneous mixture models. Due to efficient algorithms that scale well with data size and model complexity, PLEX can be used to make inferences from hundreds to thousands of taxa in only minutes on a desktop computer. It also performs probabilistic ancestral sequence reconstruction. Future versions will support detection of co-evolutionary interactions between sites, probabilistic tests of convergent evolution and rigorous testing of evolutionary hypotheses in a Bayesian framework. AVAILABILITY AND IMPLEMENTATION: PLEX v1.0 is licensed under GPL. Source code and documentation will be available for download at www.evolutionarygenomics.com/ProgramsData/PLEX. PLEX is implemented in C++ and supported on Linux, Mac OS X and other platforms supporting standard C++ compilers. Example data, control files, documentation and accessory Perl scripts are available from the website. CONTACT: David.Pollock@UCDenver.edu. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.