LSD1 is required for euchromatic origin firing and replication timing.

The chromatin-based rule governing the selection and activation of replication origins remains to be elucidated. It is believed that DNA replication initiates from open chromatin domains; thus, replication origins reside in open and active chromatin. However, we report here that lysine-specific demethylase 1 (LSD1), which biochemically catalyzes H3K4me1/2 demethylation favoring chromatin condensation, interacts with the DNA replication machinery in human cells. We find that LSD1 level peaks in early S phase, when it is required for DNA replication by facilitating origin firing in euchromatic regions. Indeed, euchromatic zones enriched in H3K4me2 are the preferred sites for the pre-replicative complex (pre-RC) binding. Remarkably, LSD1 deficiency leads to a genome-wide switch of replication from early to late. We show that LSD1-engaged DNA replication is mechanistically linked to the loading of TopBP1-Interacting Checkpoint and Replication Regulator (TICRR) onto the pre-RC and subsequent recruitment of CDC45 during origin firing. Together, these results reveal an unexpected role for LSD1 in euchromatic origin firing and replication timing, highlighting the importance of epigenetic regulation in the activation of replication origins. As selective inhibitors of LSD1 are being exploited as potential cancer therapeutics, our study supports the importance of leveraging an appropriate level of LSD1 to curb the side effects of anti-LSD1 therapy.

Pregnancy in cystic fibrosis: Review of the literature and expert recommendations.

Cystic fibrosis (CF) was historically a disease largely afflicting children. Due to therapeutic advancements, there are now more adults with CF than children. In the past decade, medications including Cystic Fibrosis Transmembrane conductance Regulator (CFTR) modulators became available that treat the underlying cause of CF and are dramatically improving lung function as well as quality and quantity of life for people with CF. As a result, more women with CF are becoming pregnant. We gathered a panel of experts in CF care, family planning, high risk obstetrics, nutrition, genetics and women with CF to review current literature on pregnancies and to provide care recommendations for this unique population.

Low Prevalence of Severe Acute Respiratory Syndrome Coronavirus 2 Among Pregnant and Postpartum Patients With Universal Screening in Seattle, Washington.

We found low prevalence of SARS-CoV-2 (2.7% [5/188]) among pregnant and postpartum patients with universal testing. Prevalence among symptomatic patients was similar under initial targeted screening (22.2% [4/18]) and universal approaches (19.1% [8/42]). Among 170 asymptomatic patients, 2 were positive or inconclusive, respectively; repeat testing at 24 hours was negative.

Development of a prenatal clinical care pathway for uncomplicated gastroschisis and literature review.

BACKGROUND: Gastroschisis is an abdominal wall defect wherein the bowel is herniated into the amniotic fluid. Controversy exists regarding optimal prenatal surveillance strategies that predict fetal well-being and help guide timing of delivery. Our objective was to develop a clinical care pathway for prenatal management of uncomplicated gastroschisis at our institution. METHODS: We performed a review of literature from January 1996 to May 2017 to evaluate prenatal ultrasound (US) markers and surveillance strategies that help determine timing of delivery and optimize outcomes in fetal gastroschisis. RESULTS: A total 63 relevant articles were identified. We found that among the US markers, intraabdominal bowel dilatation, polyhydramnios, and gastric dilatation are potentially associated with postnatal complications. Prenatal surveillance strategy with monthly US starting at 28weeks of gestational age (wGA) and twice weekly non-stress testing beginning at 32wGA is recommended to optimize fetal wellbeing. Timing of delivery should be based on obstetric indications and elective preterm delivery prior to 37wGA is not indicated. CONCLUSIONS: Close prenatal surveillance of fetal gastroschisis is necessary due to the high risk for adverse outcomes including intrauterine fetal demise in the third trimester. Decisions regarding the timing of delivery should take into consideration the additional prematurity-associated morbidity.

Prenatal Imaging Findings Predict Obstructive Fetal Airways Requiring EXIT.

OBJECTIVE: Detection of fetal airway compromise through imaging raises the possible need for ex utero intrapartum treatment (EXIT) procedures. Despite EXIT procedures involving massive resource utilization and posing increased risk to the mother, decisions for EXIT are usually based on anecdotal experience. Our objectives were to analyze prenatal consultations with potential fetal airway obstruction for imaging and obstetric findings used to determine management strategy. METHODS: Retrospective chart review was performed for prenatal abnormal fetal airway consults between 2004-2019 at a quaternary pediatric facility. Data collected included demographics, imaging characteristics, delivery information, and airway management. Our primary outcome was EXIT performance and the secondary outcome was postnatal airway management. Fisher's exact test was used to compare management decisions, outcomes, and imaging findings. RESULTS: Thirty-seven patients met inclusion criteria. The most common diagnoses observed were lymphatic malformation, teratoma, and micrognathia. Of the imaging findings collected, only midline neck mass location was associated with EXIT procedure performance. Factors associated with invasive airway support at birth were mass-induced in-utero neck extension and neck vessel compression, polyhydramnios, and micrognathia. CONCLUSIONS: Multidisciplinary input and interpretation of prenatal imaging can guide management of fetal airway-related pathology. EXIT is an overall safe procedure and can decrease risk due to airway obstruction at birth. We identified in-utero neck extension, neck vessel compression, micrognathia, and polyhydramnios as better indicators of a need for invasive airways measures at birth and suggest use of these criteria in combination with clinical judgement when recommending EXIT. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:E1357-E1362, 2021.

High-resolution profiling of human cytomegalovirus cell-free DNA in human plasma highlights its exceptionally fragmented nature.

Human cytomegalovirus (CMV) infections comprise a leading cause of newborn impairments worldwide and are pervasive concerns among the immunocompromised. Quantification of CMV viral loads is increasingly used to guide definitions of CMV disease but standardization of CMV quantitation remains problematic, mostly due to differences in qPCR amplicon sizes between clinical laboratories. Here, we used plasma cfDNA sequencing data from 2,208 samples sent for non-invasive prenatal aneuploidy screening to detect CMV and precisely measure the length of CMV fragments in human plasma. CMV reads were identified in 120 (5.4%) samples. Median cfDNA fragment size derived from CMV was significantly shorter than cfDNA derived from human chromosomes (103 vs 172 bp, p < 0.0001), corresponding to the 3rd percentile of human cfDNA. Sequencing of cfDNA from seven plasma samples from transplant patients positive for CMV confirmed the extraordinarily short nature of CMV cfDNA fragment size with a median length of 149 bp. We further show that these high-resolution measurements of CMV DNA fragment size accurately predict measured discrepancies in serum viral load measurements by different qPCR assays. These results highlight the exceptionally fragmented nature of CMV cfDNA and illustrate the promise of plasma cfDNA sequencing for quantitating viral loads through detection of fragments that would be unrecoverable by qPCR.

Gaps in Training in Interdisciplinary Social and Behavioral Sciences and Applications Among Undergraduate and Graduate Students in the 22 Top-Ranked Schools of Public Health in China.

The next generation of public health professionals requires rigorous training in behavioral health, in order to design effective behavioral interventions to respond effectively to the epidemiological transition in China. This study aimed to investigate issues in training in social and behavioral sciences in public health in China. A cross-sectional survey was conducted among 1285 and 835 last-year undergraduate and graduate public health students in 2013. The results showed that (1) majority of undergraduate students but a minority of graduate students had enrolled in psychology, social medicine, and health promotion courses; (2) very few had enrolled in other social and behavioral sciences courses; (3) high percentages of students perceived significance, needs, and interests related to social sciences courses; (4) very few were familiar with commonly used behavioral health theories and constructs, or had applied such theories/constructs to their thesis. The situation deviates from international accreditation requirement. A timely review and benchmarking are warranted.

A Paradigm Shift: Considerations in Prenatal Cell-Free DNA Screening.

BACKGROUND: Testing to determine the health of a fetus has undergone multiple iterations since the widespread adoption of amniocentesis in the 1970s, including several combinations of ultrasound and/or maternal serum screening. The clinical paradigm for prenatal screening for fetal chromosome aneuploidies was transformed by the introduction of cell-free DNA (cfDNA) screening or noninvasive prenatal screening in 2011. CONTENT: The clinical performance of cfDNA screening is well-established for the most common autosomal and sex chromosome aneuploidies with a detection rate exceeding 90% for all aneuploidies. One of the most significant advantages of cfDNA screening relative to maternal serum screening is the markedly reduced false-positive rate, which is <0.5%. The clinical implementation of cfDNA screening is discussed at length, including key biological, preanalytical, and analytical factors that affect test performance. SUMMARY: cfDNA prenatal screening for whole chromosome aneuploidies has become routine in high-risk obstetric populations. There is tremendous interest in expanding cfDNA screening to the general obstetric population. Early studies suggest that routine application of cfDNA screening is both feasible and effective, although significant economic and quality control considerations remain.

Incidence and Risk Factors of Retinopathy of Prematurity From 2 Neonatal Intensive Care Units in a Hong Kong Chinese Population.

PURPOSE: To determine the incidence and risk factors of retinopathy of prematurity (ROP) in very low birth weight (VLBW) preterm Chinese infants. DESIGN: Retrospective review. METHODS: Medical records were reviewed for all neonates screened for ROP between January 2007 and December 2012 in Hong Kong. Screening was offered to VLBW (≤1500 g) and/or preterm (gestation, ≤32 weeks) neonates using the Royal College of Ophthalmologists ROP guideline and the International Classification of ROP by 3 pediatric ophthalmologists. Maternal and neonatal covariates were analyzed using univariate and multivariable logistic regression analyses for both ROP and type 1 ROP. RESULTS: Of the 513 screened infants, the mean gestational age (GA) was 30.0 ± 2.5 weeks and the mean birth weight (BW) was 1232.6 ± 325.2 g. The incidence of ROP and type 1 ROP was 18.5% and 3.7%, respectively. In univariate analysis, a lighter BW, lower GA, blood transfusion, patent ductus arteriosus, nonsteroidal anti-inflammatory drug use, postnatal hypotension, inotropes usage, low Apgar scores, sepsis, mechanical ventilation, supplementary oxygen use, respiratory distress syndrome, anemia, surfactant use, and bronchopulmonary dysplasia were found to be associated with the development of both ROP and type 1 ROP (P < 0.05). In multivariable logistic regression analysis, BW, GA, and intraventricular hemorrhage were significant risk factors for ROP. Preeclampsia and eclampsia were the only protective factors for ROP development on multivariable logistic regression analysis (P = 0.02). CONCLUSIONS: In VLBW preterm Chinese infants, lower GA, lighter BW, and intraventricular hemorrhage were significant risk factors for ROP, whereas preeclampsia and eclampsia were protective.

Positive predictive value of non-invasive prenatal screening for fetal chromosome disorders using cell-free DNA in maternal serum: independent clinical experience of a tertiary referral center.

BACKGROUND: Non-invasive prenatal screening (NIPS) for fetal chromosome abnormalities using cell-free deoxyribonucleic acid (cfDNA) in maternal serum has significantly influenced prenatal diagnosis of fetal aneuploidies since becoming clinically available in the fall of 2011. High sensitivity and specificity have been reported in multiple publications, nearly all of which have been sponsored by the commercial performing laboratories. Once results are returned, positive and negative predictive values (PPVs, NPVs) are the performance metrics most relevant to clinical management. The purpose of this report is to present independent data on the PPVs of NIPS in actual clinical practice. METHODS: Charts were retrospectively reviewed for patients who had NIPS and were seen March 2012 to December 2013 in a tertiary academic referral center. NIPS results were compared to diagnostic genetic test results, fetal ultrasound results, and clinical phenotype/outcomes. The PPV was calculated using standard epidemiological methods. Correlation between screen results and both maternal age at delivery and gestational age at time of screening was assessed using Wilcoxon's rank sum test. RESULTS: Of 632 patients undergoing NIPS, 92 % of tests were performed in one of the four major commercial laboratories offering testing. However, all four laboratories are represented in both the normal and abnormal results groups. There were 55 abnormal NIPS results. Forty-one of 55 abnormal NIPS results were concordant with abnormal fetal outcomes, 12 were discordant, and 2 were undetermined. The PPV for all conditions included in the screen was 77.4 % (95 % CI, 63.4 - 87.3). Of 578 patients with normal NIPS results, normal pregnancy outcome was confirmed for 156 (27 %) patients. This incomplete follow-up of normal NIPS results does not affect PPV calculations, but it did preclude calculations of sensitivity, specificity, and NPV. Maternal age at delivery was significantly lower for patients with abnormal discordant results, compared to patients with abnormal concordant results (P = 0.034). Gestational age at time of screening was not associated with concordance of screen results (P = 0.722). CONCLUSIONS: The experience of using NIPS in clinical practice confirms that abnormal results cannot be considered diagnostic. Pre-test counseling should emphasize this. Diagnostic genetic testing should always be offered following abnormal NIPS results.

Successful pregnancy in patients with exstrophy-epispadias complex: A University of Washington experience.

INTRODUCTION: With advances in genitourinary reconstructive surgery, women with exstrophy-epispadias complex (EEC) have improved health and quality of life, and may reach reproductive age and consider pregnancy. Despite literature suggesting impaired fertility and higher risk with pregnancy, childbirth is possible. Medical comorbidities, including müllerian anomalies, contribute to increased risk of obstetric and urologic complications during pregnancy. OBJECTIVES: We reviewed our experience with EEC patients who achieved pregnancy to investigate (1) urological characteristics of women who achieved pregnancy; (2) pregnancy management, complications, and delivery; and (3) neonatal outcomes. We developed recommendations for managing pregnancy in women with EEC. STUDY DESIGN/RESULTS: This was a retrospective chart review of 36 female patients with EEC seen at our institution between 1996 and 2013. Female patients less than 18 years, and patients who did not have documented pregnancy were excluded. This resulted in a total of 12 patients with 22 pregnancies. All women with successful pregnancy had bladder exstrophy. The majority had undergone prior bladder augmentation (75%) and were on self-catheterization programs (92%). Thirty-six percent had symptomatic urinary tract infections (UTIs) during pregnancy. Five women had more than one pregnancy. There were four terminations of pregnancy. Of 18 desired pregnancies, there were four spontaneous abortions (SABs) (22%) and 16 live births (78%). The cesarean delivery (CD) rate was 100% (14/14), of which the majority were vertical (classical) uterine incisions with a paramedian skin incision. With the exception of one patient, there were no CD surgical complications. The mean gestational age at delivery was 36 weeks (Range 25 4/7 to 39 4/7 weeks) among eight pregnancies with known gestational age. There were no stillbirths, one neonatal death and no birth defects. DISCUSSION: Women with EEC can have successful pregnancies, though at increased risk for preterm delivery and SABs. In our cohort, the rate of SAB is similar to that described in prior studies. Symptomatic UTIs likely due to self-catheterization were common. Cesarean delivery using a paramedian skin incision and classical uterine incision were not associated with major complications in this cohort. Limitations include reliance on retrospective data and small sample size. The strength of this study is the longitudinal detailed management of pregnancies in EEC women by a single team over time. A multidisciplinary approach to providing a continuum of care from pediatrics through adolescence to adulthood optimizes successful transitions, reproductive health, and successful pregnancies. Based on our experience, an algorithm providing guidance for pregnancy management was developed.

Incidence and risk factors for retinopathy of prematurity in multiple gestations: a Chinese population study.

To determine the incidence and risk factors of retinopathy of prematurity (ROP) among new-born Chinese infants of multiple gestations.A retrospective review of medical records was performed for all neonates of multiple gestations screened for ROP between January 2007 and December 2012 in 2 neonatal intensive care units in Hong Kong. Screening was offered to very low birth weight (VLBW; ≤1500 g) and/or preterm (gestation ≤32 weeks) neonates using the Royal College of Ophthalmologists ROP guideline and the International Classification of ROP by 3 pediatric ophthalmologists. Maternal and neonatal covariates were analyzed using univariate and multivariate regression analyses for both ROP and Type 1 ROP.A total of 153 Chinese infants of multiple gestations were included in the study. The mean gestational age (GA) was 30.8 ±â€Š2.4 weeks and the mean birth weight (BW) was 1284.8 ±â€Š267.4 g. The incidence of ROP and Type 1 ROP was 11.8% and 3.9%, respectively. On univariate analysis, younger GA, lighter birth weight, postnatal hypotension, inotropes use, bronchopulmonary disease, and intraventricular hemorrhage were common independent risk factors for the development of ROP and Type 1 ROP (all P ≤ 0.04). On multivariate analysis, younger GA, surfactant use, invasive mechanical ventilation, higher mean oxygen concentration, thrombocytopenia, intraventricular hemorrhage, total parental nutrition, and hypoglycemia were significant risk factors for ROP. For Type 1 ROP, there were no significant dependent risk factors.In preterm Chinese infants born from multiple gestations, prematurity, lighter weight, postnatal hypotension, inotropes use, bronchopulmonary dysplasia, and an intraventricular hemorrhage were common independent risk factors for the development of ROP and Type 1 ROP.

Differences in risk factors for retinopathy of prematurity development in paired twins: a Chinese population study.

PURPOSE: To determine the differences in risk factors for retinopathy of prematurity (ROP) in paired twins. METHODS: A retrospective medical record review was performed for all paired twins screened for ROP between 2007 and 2012. Screening was offered to very low birth weight (≤ 1500 grams) and preterm (≤ 32 weeks) neonates. Twins 1 and 2 were categorized based on the order of delivery. Maternal and neonatal covariates were analyzed using univariate and multivariate regression analyses for both ROP and Type 1 ROP. RESULTS: In 34 pairs of Chinese twins, the mean gestational age (GA) was 30.2 ± 2.0 weeks. In Twin 1, smaller GA (OR = 0.44, P = 0.02), higher mean oxygen concentration (OR = 1.34, P = 0.03), presence of thrombocytopenia (OR = 1429.60, P < 0.0001), and intraventricular hemorrhage (OR = 18.67, P = 0.03) were significant risk factors for ROP. For Twin 2, a smaller GA (OR = 0.45, P = 0.03) was the only risk factor. There were no significant risk factors for ROP in Twin 1 or Twin 2 on multivariate analysis. CONCLUSION: In Chinese twin pairs, smaller GA was the only common risk factor for ROP while Twin 1 was more susceptible to the postnatal risks for ROP.

Successful induction of labor of late-second-trimester conjoined twins: an alternative to hysterotomy.

BACKGROUND: To demonstrate that vaginal delivery is a safe alternative to hysterotomy when planning pregnancy termination of late-second-trimester conjoined twins. We present two cases of conjoined twins in the late second trimester desiring pregnancy termination. CASE: The first case involved a multiparous 29-year-old woman at 23 6/7 weeks of gestation with thoraco-omphalopagus conjoined twins. The second case involved an 18-year-old primigravid woman at 25 1/7 weeks of gestation with pyopagus conjoined twins. Both desired pregnancy termination and to avoid hysterotomy. Inductions were initiated with Laminaria and augmented with vaginal misoprostol or oxytocin. Both patients had uncomplicated vaginal deliveries of intact conjoined twins without significant maternity morbidity. CONCLUSION: Induction of labor and vaginal delivery can be a safe method for late-second-trimester termination of pregnancies with conjoined twins.

PRImary care Streptococcal Management (PRISM) study: identifying clinical variables associated with Lancefield group A ß-haemolytic streptococci and Lancefield non-Group A streptococcal throat infections from two cohorts of patients presenting with an acute sore throat.

OBJECTIVE: To assess the association between features of acute sore throat and the growth of streptococci from culturing a throat swab. DESIGN: Diagnostic cohort. SETTING: UK general practices. PARTICIPANTS: Patients aged 5 or over presenting with an acute sore throat. Patients were recruited for a second cohort (cohort 2, n=517) consecutively after the first (cohort 1, n=606) from similar practices. MAIN OUTCOME: Predictors of the presence of Lancefield A/C/G streptococci. RESULTS: The clinical score developed from cohort 1 had poor discrimination in cohort 2 (bootstrapped estimate of area under the receiver operator characteristic (ROC) curve (0.65), due to the poor validity of the individual items in the second data set. Variables significant in multivariate analysis in both cohorts were rapid attendance (prior duration 3 days or less; multivariate adjusted OR 1.92 cohort, 1.67 cohort 2); fever in the last 24 h (1.69, 2.40); and doctor assessment of severity (severely inflamed pharynx/tonsils (2.28, 2.29)). The absence of coryza or cough and purulent tonsils were significant in univariate analysis in both cohorts and in multivariate analysis in one cohort. A five-item score based on Fever, Purulence, Attend rapidly (3 days or less), severely Inflamed tonsils and No cough or coryza (FeverPAIN) had moderate predictive value (bootstrapped area under the ROC curve 0.73 cohort 1, 0.71 cohort 2) and identified a substantial number of participants at low risk of streptococcal infection (38% in cohort 1, 36% in cohort 2 scored ≤1, associated with a streptococcal percentage of 13% and 18%, respectively). A Centor score of ≤1 identified 23% and 26% of participants with streptococcal percentages of 10% and 28%, respectively. CONCLUSIONS: Items widely used to help identify streptococcal sore throat may not be the most consistent. A modified clinical scoring system (FeverPAIN) which requires further validation may be clinically helpful in identifying individuals who are unlikely to have major pathogenic streptococci.

Germline mosaicism does not explain the maternal age effect on trisomy.

A variety of hypotheses have been proposed to explain the association between trisomy and increasing maternal age in humans, virtually all of which assume that the underlying mechanisms involve meiotic errors. However, recently Hultén and colleagues [Hulten et al., 2010b] proposed a provocative model-the Oocyte Mosaicism Selection Model (OMSM)-that links age-dependent trisomy 21 to pre-meiotic errors in the ovary. Specifically, they propose that nondisjunctional events occur in a proportion of germ cells as they mitotically proliferate, resulting in mosaicism for trisomy 21. Assuming that the presence of an additional chromosome 21 delays meiotic progression, these cells would be ovulated later in reproductive life, resulting in an age-dependent increase in aneuploid eggs. Because this model has important clinical implications, we initiated studies to test it. We first analyzed oocytes from two trisomy 21 fetuses, combining immunostaining with FISH to determine the likelihood of detecting the additional chromosome 21 at different stages of meiosis. The detection of trisomy was enhanced during the earliest stage of prophase (leptotene), before homologs synapsed. Accordingly, in subsequent studies we examined the chromosome content of leptotene oocytes in seven second trimester female fetuses, analyzing three chromosomes commonly associated with human trisomies (i.e., 13, 16, and 21). In contrast to the prediction of the OMSM, we found no evidence of trisomy mosaicism for any chromosome. We conclude that errors in pre-meiotic germ cells are not a major contributor to human aneuploidy and do not provide an explanation for the age-related increase in trisomic conceptions.

Genome rearrangements caused by depletion of essential DNA replication proteins in Saccharomyces cerevisiae.

Genetic screens of the collection of ~4500 deletion mutants in Saccharomyces cerevisiae have identified the cohort of nonessential genes that promote maintenance of genome integrity. Here we probe the role of essential genes needed for genome stability. To this end, we screened 217 tetracycline-regulated promoter alleles of essential genes and identified 47 genes whose depletion results in spontaneous DNA damage. We further showed that 92 of these 217 essential genes have a role in suppressing chromosome rearrangements. We identified a core set of 15 genes involved in DNA replication that are critical in preventing both spontaneous DNA damage and genome rearrangements. Mapping, classification, and analysis of rearrangement breakpoints indicated that yeast fragile sites, Ty retrotransposons, tRNA genes, early origins of replication, and replication termination sites are common features at breakpoints when essential replication genes that suppress chromosome rearrangements are downregulated. We propose mechanisms by which depletion of essential replication proteins can lead to double-stranded DNA breaks near these features, which are subsequently repaired by homologous recombination at repeated elements.

Ambiguous genitalia: what prenatal genetic testing is practical?

Concern for ambiguous genitalia or chromosome-phenotype discordance detected in a prenatal setting has increased over the last two decades. Practitioners faced with this prenatal finding have a variety of genetic tests available to them; however, it is unclear to what extent prenatal testing for disorders of sex development (DSD) is useful or practical. We undertook a retrospective review of the medical records of 140 individuals evaluated through the DSD clinic at Seattle Children's Hospital with birthdates from 01/01/1994 through 08/16/2011 to determine the rate of prenatal detection of ambiguous genitalia in individuals with DSD, what prenatal diagnostic workup was undertaken, and the postnatal outcome, including whether a postnatal genetic diagnosis was confirmed. Of all 140 subjects, 34 (24%) were identified prenatally. The most common postnatal diagnoses were penoscrotal hypospadias with transposition of the scrotum with no known genetic cause (24/140; 17%) and 21-hydroxylase deficiency (20/140; 14%). Apart from these, no single diagnosis comprised more than a few cases. Prenatal diagnostic testing varied widely, from no tests to multiple molecular tests with amniotic fluid hormone concentrations. In the absence of other fetal anomalies or growth retardation on ultrasound, prenatal karyotype with fluorescence in situ hybridization for the SRY gene is the most useful test when ambiguous genitalia is suspected. Further prenatal testing for Smith-Lemli-Opitz syndrome in 46,XY individuals and congenital adrenal hyperplasia in 46,XX individuals may be considered. However, targeted molecular testing for rare DSD conditions in the absence of a family history of DSD has a low yield.