BACKGROUND: Early identification of fetal gender is crucial for managing gender-linked genetic disorders. This study aimed to evaluate the predictive performance of anogenital distance (AGD) and genital tubercle angle (GTA) for fetal sex determination during the first trimester.
METHODS: A multicenter retrospective cohort study was conducted on 312 fetal cases between 11 and 13 + 6 weeks of gestation from two tertiary hospitals. AGD and GTA measurements were taken from midsagittal plane images using ultrasound, with intra- and inter-reader reproducibility assessed. Binomial logistic regression and ROC curve analysis were employed to determine the diagnostic performance and optimal cutoff points.
RESULTS: AGD had a mean of 7.16 mm in male fetuses and 4.42 mm in female fetuses, with a sensitivity of 88.8%, specificity of 94.4%, and an area under the ROC curve (AUC) of 0.931 (95% CI: 0.899-0.962) using 5.74 mm as a cutoff point. For GTA, the mean was 35.90 degrees in males and 21.57 degrees in females, with a sensitivity of 92%, specificity of 84.7%, and an AUC of 0.932 (95% CI: 0.904-0.961) using 28.32 degrees as a cutoff point. The reproducibility results were excellent for AGD (intra-operator ICC = 0.938, inter-operator ICC = 0.871) and moderate for GTA (intra-operator ICC = 0.895, inter-operator ICC = 0.695).
CONCLUSIONS: The findings suggest that AGD and GTA are reliable markers for early fetal sex determination, with AGD showing higher reproducibility. The findings highlight the feasibility and accuracy of these non-invasive sonographic markers and their potential usefulness in guiding timely interventions and enhancing the management of gender-linked genetic conditions.

The timing of exposure to the steroid hormone, testosterone, produces activational and organizational effects in vertebrates. These activational and organizational effects are hypothesized to relate with the number of female mating partners and reproductive success in males. We tested this hypothesis by examining 151 wild degu (Octodon degus) males across a 10-year study. We quantified the association between adult serum testosterone levels (i.e., an indirect index of adult activational effects) and anogenital distance (AGD) length (i.e., a direct index of fetal organizational effects), and their interaction on the number of female mating partners and reproductive success. We found no evidence of an association between adult male serum testosterone levels and the number of female mating partners, or between adult male serum testosterone levels and reproductive success. However, male AGD was positively associated with reproductive success, but not so with the number of female mating partners. Additionally, the positive association between male AGD and male reproductive success was mediated by the number of mates. Our findings do not support major roles of activational or organizational effects of testosterone on the number of female mating partners and its consequences on male reproductive success. Instead, our results suggest that compared with individual male attributes, the female social environment plays a more important role in driving male reproductive success.

UNASSIGNED: Assessment of genitalia is an important part of the neonatal examination. Regional, racial, and ethnic variations in phallic length have been documented. Clitoral dimensions may also show similar variations. Normal values for neonatal clitoral measurements may help the pediatrician/neonatologist to accurately diagnose clitoromegaly and underlying etiology to guide appropriate investigations. Data on clitoral measurements is limited with only one study from India (Kolkata). Hence we aimed in this study to generate south Indian data on neonatal clitoral dimensions, anogenital distance, and anogenital ratio. Two hundred and fifty two hemodynamically stable term female neonates without ambiguous genitalia/vulval masses born in a community hospital. Hospital-based cross-sectional study. 1. To measure clitoral dimensions, anogenital distance, and anogenital ratio in female neonates. 2. To correlate clitoral dimensions, anogenital distance, and ratio with anthropometric measurements, gestational age, and maternal comorbidities.
UNASSIGNED: Measurements were recorded using a digital vernier caliper, under strict aseptic precautions with labia majora gently separated and the baby held in a frog-leg position. Clitoral length, width, and anogenital distance were measured and anogenital ratio and clitoral index were calculated.
UNASSIGNED: In term neonates, the mean ± SD of clitoral length (CL), clitoral width (CW), anogenital ratio (AGR) were 6.34 ± 1.75 mm, 6.39 ± 1.27 mm and 0.39 ± 0.05, respectively. The 3rd and 97th centiles for mean clitoral length were 3.55 and 9.93 mm, for mean clitoral width were 3.37 and 8.35 mm, and for AGR were 0.28 and 0.48, respectively. These clitoral dimensions in south Indian neonates were higher than those from East India (Kolkata), lower than Nigerian babies, and similar to Israeli neonates. Mean CL and CW had no statistical correlation with birth weight, gestational age, head circumference, or length in term neonates. A significant correlation was noted between pregnancy-induced hypertension and mean clitoral width, and between gestational diabetes and AGR.
UNASSIGNED: Normative values for clitoral dimensions (length and width) and AGR for south Indian term female neonates have been established. 97th centiles of 9.93 mm (CL) and 8.35 mm (CW) and 0.48 mm (AGR) may be used as practical cut-offs to diagnose clitoromegaly and virilization. Clitoral measurements had no statistical correlation with birth weight, gestational age, or anthropometry. Clitoral measurements exhibit ethnic and racial differences, thus emphasizing the importance of regional cut-offs and need for more studies from different parts of India on CL.

BACKGROUND: Measurement of rat anogenital distance (AGD) dates to at least 1912. Increased interest in endocrine disrupting chemicals and the use of AGD as a biomarker for fetal androgen effects have increased the number of studies with this endpoint in recent decades. A literature review revealed different landmarks, methods of measurement, and methods to adjust for body weight differences. AGD is often reported to hundredths of millimeters and as such, deserves precision in all these aspects. This paper presents recommendations for the measurement and analysis of rodent AGD.
METHODS: Literature and regulatory guidance documents that mentioned or measured rodent AGD were reviewed. Four adjustment methods were evaluated using available online data from three rat studies each with two generations of offspring.
RESULTS: Tabulation of studies reveals that species/stocks and time of data collection, but more importantly anatomical landmarks and methods of measurement have produced a variety of results which are difficult to compare. Not all studies have adjusted for test article effects on body weight (and thus size). The four adjustment methods were fairly comparable.
CONCLUSIONS: Recommendations are as follows. A microscopic method should be used to measure AGD of late rodent fetuses and early postnatal pups. The caudal edge of the genital tubercle and the cranial edge of the anus are clear and identifiable landmarks. The simplest adjustment is to divide individual AGDs by the cube root of animals\' body weight. These recommendations will help ensure data consistency and accuracy, and facilitate meaningful comparisons across laboratories and chemical classes.

BACKGROUND: The anogenital distance (AGD) is considered a postnatal readout of early fetal androgen action. Little is known of prenatal AGD and how it correlates with AGD postnatally.
OBJECTIVE: We present longitudinal measurements of fetal- and infant AGD. We evaluate the impact of testosterone and dihydrotestosterone at minipuberty on AGD and penile size.
METHODS: Secondary analyses of an observational, prospective pregnancy and birth cohort, COPANA (2020-2022).
METHODS: Copenhagen University Hospital - Rigshospitalet.
METHODS: 685 healthy, singleton pregnant women enrolled, 657 women attended 3rd trimester ultrasound, 589 infants completed follow-up.
METHODS: 3rd trimester ultrasound (GW29-34): Fetal AGD. Minipuberty clinical examination (app. 3.5 months postpartum): infant AGD, penile width and stretched length (SPL), circulating testosterone and dihydrotestosterone (LC-MS/MS).
RESULTS: AGD was available in 650/657 fetuses (310 boys) and 588/589 infants (287 boys). Boys had longer fetal and infant AGD compared to girls; fetal AGDas: mean (SD) 21.4 mm (±3.5), fetal AGDaf: 12.8 mm (±2.3), p < 0.001, infant AGDas: 32.0 mm (±5.6) and infant AGDaf: 15.8 (±3.3), p < 0.001. Fetal AGD correlated with infant AGD in boys and girls (Spearman\'s r = 0.275, p < 0.001 and r = 0.189, p = 0.001 respectively), but not with circulating testosterone or dihydrotestosterone at minipuberty. Penile size correlated positively with circulating androgen levels at minipuberty, i.e.: SPL vs testosterone: r = 0.235, p < 0.001.
CONCLUSIONS: AGD is sexual dimorphic already in the 3rd trimester. Fetal and infant AGD correlates. AGD is associated with body size but not circulating androgen levels at minipuberty. These findings suggest that fetal and infant AGD, reflect androgen action during early fetal development.

UNASSIGNED: To describe the anatomical abnormalities of hypospadias before puberty using current commonly used anthropometric index data and predict postoperative diagnostic classification.
UNASSIGNED: Children with hypospadias before puberty who were initially treated at Sichuan Provincial People\'s Hospital from April 2021 to September 2022 were selected. We recorded their preoperative penoscrotal distance, anogenital distance, 2D:4D finger ratio, and postoperative hypospadias classification. The receiver operating character curve was used for univariate analysis of the diagnostic predictive value of each index for hypospadias classification in the training set. Binary logistic regression, random forest, and support vector machine models were constructed. In addition, we also prospectively collected data from October 2022 to September 2023 as a test set to verify the constructed machine learning models.
UNASSIGNED: This study included 389 cases, with 50 distal, 167 midshaft, and 172 proximal cases. In the validation set, the sensitivity of the binary LR, RF, and SVM was 17%, 17% and 0% for identifying the distal type, 61%, 55% and 64% for identifying the midshaft type, and 56%, 60% and 48% for identifying the proximal type, respectively. The sensitivity of the three-classification RF and SVM models was 17% and 17% for distal type, 64% and 73% for midshaft type, 60% and 60% for proximal type, respectively. In the Testing set, the sensitivity of the binary LR, RF and SVM was 6%, 0% and 0% for identifying the distal type, 64%, 55% and 66% for identifying the midshaft type, and 48%, 62% and 39% for identifying the proximal type, respectively. The sensitivity of the three-classification RF and SVM models was 12% and 0% for distal type, 57% and 77% for midshaft type, and 65% and 53% for proximal type, respectively. Compared with binary classification models, the sensitivity of the three-classification models for distal type was not improved.
UNASSIGNED: Anogenital distance and penoscrotal distance have a favorable predictive value for midshaft and proximal hypospadias, among which AGD2, with higher test efficiency and stability, is recommended as the preferred anogenital distance indicator. The 2D:4D finger ratio (RadioL, RadioR) has little predictive value for hypospadias classification.

Dibutyl phthalate (DBP) and di(2-ethylhexyl) phthalate (DEHP), two common types of phthalates, are known to cause reproductive and developmental toxicity in animals and humans. The reference doses (RfD) of DBP and DEHP should be determined by sensitive endpoints. We here aimed to identify sensitive endpoints for DBP- and DEHP-induced such toxicity using published literatures. By examining the impacts of maternal exposure to DBP or DEHP on anogenital distance (AGD) and semen quality of offspring, we discovered that DBP or DEHP caused AGD decline in boys but increase in girls with DBP being more potent and the first 14weeks of pregnancy being more susceptible, suggesting a chemical- and time-dependent phenomenon. We also identified AGD shortening and total sperm count reduction as two sensitive endpoints for DBP- or DEHP-induced reproductive and developmental toxicity, respectively. Based upon these two endpoints and the employment of the Bayesian benchmark dose approach with an uncertainty factor of 3,000, we estimated the RfD values of DBP and DEHP were 15 μg/kg/day and 36 μg/kg/day, respectively. Thus, we uncover previously unrecognized phenomena of DBP- or DEHP-induced reproductive and developmental toxicity and establish new and comparable or more conservative RfDs for the risk assessment of phthalates exposure in humans.

OBJECTIVE: Do different boys with different types of cryptorchidism exhibit different anogenital distances (AGDs)?
CONCLUSIONS: Length of AGD seemed to differ in different groups of patients with cryptorchidism.
BACKGROUND: AGD, which is used as an indicator of prenatal androgen action, tends to be shorter in boys with cryptorchidism compared to unaffected boys. Shorter AGDs have also been reported in boys with hypospadias, in men with poor semen quality, and in men with testicular cancer.
METHODS: A prospective descriptive cohort study was performed using data from consecutively selected boys with cryptorchidism (n = 169) operated in a single center over a period of 3 years (September 2019 to October 2022).
METHODS: AGD was measured in 169 infant boys, at 3 to 26 months of age, during anesthesia with a vernier caliper measuring the distance from the anus to the base of the scrotum (AGDAS) and from the anus to the anterior base of the penis (AGDAP) in two body positions according to the methods by \'The Infant Development and the Environment Study\' (TIDES) and \'Cambridge Baby Growth Study\', resulting in four mean values per patient (TIDES AGDAS/AP and Cambridge AGDAS/AP). Normal values for AGD by age were set by our hospital Department of Growth and Reproduction based on a large cohort of healthy infant boys (n = 1940). Testicular biopsies were performed at orchidopexy as a clinical routine. The germ cell number (G/T) and type Ad spermatogonia number (AdS/T) per cross-sectional tubule of at least 100 and 250 tubules, respectively were measured and related to normal samples. Blood samples were obtained by venipuncture for measuring serum LH, FSH, and inhibin B. They were analyzed in our hospital Department of Growth and Reproduction where the normal reference was also established. Correlations between the four mean AGD measurements for each boy were evaluated by Spearman rank correlation analyses. The AGD measurement of every boy was transferred to the multiple of the median (MoM) of the normal AGD for age and named MoM AGD.
RESULTS: There were 104 boysoperated for unilateral, and 47 boys operated for bilateral, undescended testes, whereas 18 boys had vanished testis including one boy with bilateral vanished testes. Only 6% of cases with vanished testes had a MoM AGD higher than the normal median compared to 32% with undescended testes (P < 0.05). MoM AGD increased with the age at surgery for boys with vanished testis (Spearman r = 0.44), but not for boys with undescended testes (Spearman r = 0.14). Boys with bilateral cryptorchidism had longer AGDs and more often had hypogonadotropic hypogonadism than boys with unilateral cryptorchidism (P < 0.005) and (P < 0.000001).
CONCLUSIONS: Although being the largest published material of AGD measurements of infant boys with cryptorchidism, one limitation of this study covers the quite small number of patients in the different groups, which may decrease the statistical power. Another limitation involves the sparse normal reference material on G/T and AdS/T. Finally, there are currently no longitudinal studies evaluating AGD from birth to adulthood and evaluating childhood AGD in relation to fertility outcome. Our study is hypothesis generating and therefore the interpretation of the results should be regarded as exploratory rather than reaching definite conclusions.
CONCLUSIONS: The study findings are in agreement with literature as the total included group of boys with cryptorchidism exhibited shorter than normal AGDs. However, new insights were demonstrated. Boys with vanished testis had shorter AGDs compared to unaffected boys and to boys with undescended testes. This finding challenges the current concept of AGD being determined in \'the masculinization programming window\' in Week 8 to 14 of gestation. Furthermore, boys with bilateral cryptorchidism had longer AGDs and more often had hypogonadotropic hypogonadism than boys with unilateral cryptorchidism, suggesting that the lack of fetal androgen in hypogonadotropic hypogonadism is not that significant.
BACKGROUND: No external funding was used and no competing interests are declared.
BACKGROUND: The trial was not registered in an ICMJE-recognized trial registry.

OBJECTIVE: To investigate the anogenital distance from the upper verge of the anus to the posterior fourchette (AGDAF), FASL, and BCL2 combination as a reliable and non-invasive tool for the diagnosis of endometriosis.
METHODS: This study included 100 women with endometriosis and 50 women without endometriosis as the control group. All cases underwent history taking, body mass index (BMI) measurement, AGD measurement, and FASL and BCL2 immunohistochemical staining of the eutopic endometrial tissue.
RESULTS: This study included 150 women divided into endometriosis and control groups. Endometriosis cases significantly had shorter AGDAF, 22.9 ± 2.6 mm, compared with the control group, 27.3 ± 3.5 mm (P < 0.001). Lower FASL and higher BCL2 expression were associated with endometriosis (P < 0.001). The combined measurement of AGDAF (cut-off point 24.55 mm) with FASL and BCL2 was associated with endometriosis (P < 0.001). The combined diagnostic sensitivity, specificity, positive predictive value, and negative predictive value of AGDAF, FASL, and BCL2 were 83%, 78%, 87.3%, and 69.6%, respectively. The area under the curve was greater for AGDAF, FASL, and BCL2 in combination than for individual measurements.
CONCLUSIONS: Combining short AGDAF with high BCL2 and low FASL is a highly sensitive, non-invasive diagnostic tool for endometriosis.

Per- and polyfluoroalkyl substances (PFAS) are persistent organic pollutants ubiquitous in the environment and humans. In-utero PFAS exposure is associated with numerous adverse health impacts. However, little is known about how prenatal PFAS mixture exposure affects offspring\'s neurobehavioral function. This study aims to determine the causal relationship between in-utero PFAS mixture exposure and neurobehavioral changes in Sprague-Dawley rat offspring. Dams were exposed via drinking water to the vehicle (control), an environmentally relevant PFAS mixture, or a high-dose PFAS mixture. The environmentally relevant mixture was formulated to resemble measured tap water levels in Pittsboro, NC, USA (10 PFAS compounds; sum PFAS =758.6 ng/L). The high-dose PFAS load was 3.8 mg/L (5000×), within the range of exposures in the experimental literature. Exposure occurred seven days before mating until birth. Following exposure to PFAS-laden water or the vehicle during fetal development, neurobehavioral toxicity was assessed in male and female offspring with a battery of motor, cognitive, and affective function tests as juveniles, adolescents, and adults. Just before weaning, the environmentally relevant exposure group had smaller anogenital distances compared to the vehicle and high-dose groups on day 17, and males in the environmentally relevant exposure group demonstrated lower weights than the high-dose group on day 21 (p < 0.05). Reflex development delays were seen in negative geotaxis acquisition for both exposure groups compared to vehicle-exposed controls (p = 0.009). Our post-weaning behavioral measures of anxiety, depression, and memory were not found to be affected by maternal PFAS exposure. In adolescence (week five) and adulthood (week eight), the high PFAS dose significantly attenuated typical sex differences in locomotor activity. Maternal exposure to an environmentally relevant PFAS mixture produced developmental delays in the domains of pup weight, anogenital distance, and reflex acquisition for rat offspring. The high-dose PFAS exposure significantly decreased typical sex differences in locomotor activity.