Evaluation of the infant with atypical genital appearance (difference of sex development)

INTRODUCTION — Infants born with genitals that do not appear typically male or female, or that have an appearance discordant with the chromosomal sex (picture 1), are classified as having a difference (or disorder) of sex development (DSD; terminology discussed further below). DSDs with a genital appearance that is sufficiently atypical to prompt evaluation occur in approximately 1 in 1000 to 4500 live births [1-4].

The evaluation of the infant with a DSD is presented here. Specific causes of DSDs and the management of affected infants are discussed separately. (See "Causes of differences of sex development" and "Management of the infant with atypical genital appearance (difference of sex development)".)

TERMINOLOGY

Difference of sex development — The diagnostic category "disorder of sex development (DSD)" was proposed in a 2006 consensus conference, replacing potentially pejorative terms including "pseudohermaphrodite," "hermaphrodite," and "intersex" [5]. Since then, many groups have modified the term to "difference of sex development" in response to concerns that "disorder" is unnecessarily negative [4,6].

Concerns have been raised that DSD is an overly broad term that could be interpreted to encompass conditions in which no issues with genital appearance or gender identity are expected; and also that a broad umbrella term like DSD is unnecessary because it lacks sufficient specificity to be helpful diagnostically. Many patient support groups and advocacy groups do not accept the DSD designation and feel that it should be abandoned by the medical community.

Until a consensus is reached on this issue, we will use the term DSD but only apply it to patients with atypical development of the external genitalia. Many of these individuals present as newborns with an atypical genital appearance sometimes termed "ambiguous genitalia." We will not use the term to refer to conditions in which genital/gender discordance is not expected, such as Klinefelter syndrome, Turner syndrome, undescended testes, or isolated and nonsevere (standard) forms of hypospadias [7]. (See 'Indications for evaluation' below.)

Glossary of other terms — The vocabulary used to describe features of DSDs can also be confusing and is sometimes inconsistently applied. A glossary of terms is provided (table 1).

TYPICAL SEX DEVELOPMENT AND ITS VARIATIONS — A logical approach to the infant with a DSD requires an understanding of typical human sex development. This process is reviewed here briefly and discussed in detail elsewhere. (See "Typical sex development".)

●Gonadal differentiation – The ovaries and testes derive from a primitive bipotential gonad. In typical XY individuals, expression of the SRY gene (sex-determining region on the Y chromosome) activates pathways that cause the gonads to differentiate as testes. In typical XX individuals, the absence of SRY and active repression of testicular pathways causes the gonads to differentiate into ovaries. Individuals with DSDs may have complete absence of gonadal development (complete gonadal dysgenesis), partial but incomplete gonadal development (partial gonadal dysgenesis), a gonad atypical for chromosomal sex (eg, XX testicular DSD), or a gonad with both ovarian and testicular features (an ovotestis).

●Gonadal function – The Leydig cells of the testis produce testosterone, which is converted to dihydrotestosterone by the enzyme 5-alpha-reductase type 2, which is expressed outside of the testis. Dihydrotestosterone and, to a lesser extent, testosterone activate the androgen receptor. The Leydig cells also produce insulin-like 3 (INSL3). The Sertoli cells of the testis produce anti-müllerian hormone (AMH; also known as müllerian-inhibiting substance [MIS] and müllerian regression factor). During embryonic and fetal development, the ovary does not produce significant amounts of testosterone, INSL3, or AMH. Individuals with DSDs may produce testosterone, dihydrotestosterone, INSL3, and AMH to varying degrees, depending on the underlying condition.

●Gonadal position – In typical XY individuals, INSL3 and testosterone cause the gonads to descend into the scrotum. In typical XX individuals, these factors are absent and the gonads remain in the abdomen. Individuals with DSDs may have gonads located in the abdomen, the inguinal region, or the labioscrotal folds.

●Development of the labioscrotal folds – The embryonic genital swellings give rise to the labioscrotal folds. In typical XY individuals, the action of testosterone and dihydrotestosterone cause the labioscrotal folds to fuse to form the scrotum and become rugated (wrinkled) and pigmented. In typical XX individuals, the labioscrotal folds remain unfused and develop into the labia majora. Individuals with DSDs may have varying degrees of fusion, rugation, and pigmentation of the labioscrotal folds, depending on the degree of androgen action.

●Size and shape of the clitorophallus – In typical XY individuals, the action of testosterone and dihydrotestosterone cause the genital tubercle to increase in size and straighten to form the penis (figure 1). In typical XX individuals, the genital tubercle develops into the clitoris. Individuals with DSDs may have varying degrees of size and curvature of the clitorophallus, depending on the degree of androgen action.

●Location of the urethra – In typical XY individuals, androgen action causes the urethral plate to contribute to the penile urethra. The urethra within the glans penis forms by a complex process of tissue remodeling and canalization. The result is a urethral meatus (orifice) located at the tip of the glans penis. In typical XX individuals, the absence of androgen action allows the separation of the urinary and reproductive tracts and the development of distinct urethral and vaginal orifices. Individuals with DSDs may have a urethral meatus located anywhere between the tip of the glans and the perineum. Of note, some individuals may have a blind-ending dimple at the tip of the clitorophallus that may be mistaken for the urethral meatus. Partial androgen action may result in incomplete separation of the urinary and reproductive tracts, leading to a common urogenital sinus with a single orifice rather than separate urethral and vaginal tracts with distinct orifices.

●Internal reproductive structures – In typical XY individuals, local testosterone secretion causes ipsilateral development of the Wolffian ducts into the epididymis, vas deferens, seminal vesicle, and ejaculatory duct (figure 2) and local AMH secretion leads to ipsilateral regression of the müllerian ducts. In typical XX individuals, the absence of androgen action results in regression of the Wolffian ducts and the absence of AMH leads to retention of the müllerian ducts, which develop into the uterus, Fallopian tubes, and upper two-thirds of the vagina. The lower portion of the vagina forms from urogenital sinus epithelium without müllerian or Wolffian input. Individuals with DSD may have varying degrees of development of Wolffian and müllerian structures, and asymmetric gonadal function may result in asymmetry of the internal reproductive structures.

OVERVIEW OF THE EVALUATION

Guiding principles — Clinicians managing the initial care and evaluation of an infant with a suspected DSD should:

●Determine an accurate diagnosis when possible.

●Avoid designating gender/sex, naming the child, or completing the birth certificate prematurely.

●Prevent the salt-wasting crisis that can occur with certain types of congenital adrenal hyperplasia (CAH).

●Expedite a thorough workup to guide gender/sex designation (and, in particular, assessing the sex chromosome complement as quickly as possible), explaining the importance of this workup to families and acknowledging the distress that may be present.

●Place the DSD in an appropriate context (ie, the overall health of the baby). If the baby is healthy, this should be emphasized in discussions with the family. If the baby has health concerns, evaluation and management of those concerns may assume a higher priority than the suspected DSD.

●Present the DSD as a chronic issue that affects multiple systems and may require lifelong management and support. Avoid presenting the DSD as a strictly anatomic issue that can be "corrected" by surgery.

●Consult with specialist teams and/or transfer the patient to a higher-level facility if appropriate and possible.

Indications for evaluation

●DSD evaluation recommended – The possibility of a DSD should be considered in any infant with frankly atypical genital appearance and also in those who may have less obvious presentations, such as:

•Generally male genital appearance with any of the following features:

-Bilaterally nonpalpable gonads (picture 2)

-Severe hypospadias (scrotal or perineal ectopic meatus, severe penile curvature, fusion of the foreskin with the scrotum, and/or a small glans size [<14 mm before one year of age]) (picture 3) with descended testes. Approximately one-half of such infants have clinically relevant variants in DSD-associated genes [7,8].

-Any degree of hypospadias accompanied by unilateral or bilateral cryptorchidism (nonpalpable gonad) and/or micropenis (stretched penile length less than 2.5 cm in a full-term infant) (picture 4A-B)

-Genital appearance discordant with the sex chromosomes

•Generally female genital appearance with:

-Clitoromegaly – Clitoral width >6 mm or clitoral length >9 mm (picture 5A-B)

-Posterior labial fusion – Anogenital ratio >0.5 (figure 3); single opening (common urogenital sinus) instead of a separate opening for the urethra and vagina (picture 6)

-Gonads palpable in the labioscrotal folds or the inguinal region

-Genital appearance discordant with the sex chromosomes (picture 1)

Detailed descriptions of physical manifestations of DSDs are given below. (See 'Physical examination' below.)

●DSD evaluation not required – Although DSDs can produce a range of phenotypes, a comprehensive evaluation is not generally recommended for the following presentations, because the yield of such a comprehensive DSD evaluation is thought to be low, based on clinical experience:

•Male-appearing genitals with standard hypospadias (ectopic meatus on the glans, penile shaft and/or penoscrotal junction, with a normal-sized phallus and glans [>13 mm in the first year of life] and penile curvature that is no more than mild to moderate) if both gonads are palpable and there is no micropenis. While some DSD may present with this appearance, most individuals with this presentation do not have an underlying DSD [7-11].

•Male-appearing genitals with isolated micropenis (though such infants should be evaluated for hypogonadotropic hypogonadism/hypopituitarism).

•Female-appearing genitals with an atypical clitoral hood but no clitoromegaly.

(See "Clinical features and diagnosis of male hypogonadism" and "Hypospadias: Pathogenesis, diagnosis, and evaluation".)

Diagnostic approach — The diagnostic approach involves each of the following steps, as summarized in the table (table 2):

●Prompt testing for CAH – For all infants with bilateral nonpalpable gonads and suspected DSD, evaluate for CAH by measuring 17-hydroxyprogesterone (17-OHP) and, if possible, other adrenal hormones and precursors (eg, androstenedione, 17-hydroxypregnenolone, and 11-deoxycortisol, which are sometimes available as a CAH panel). Evaluation of adrenal function may require a stimulation test using adrenocorticotropic hormone (ACTH). CAH is the most common cause of atypical genital appearance in infants and presents with varying degrees of virilization. It is important to diagnose and treat CAH promptly because of the associated risk for adrenal crisis, which can occur as early as five days of life [12]. (See 'Initial laboratory testing' below.)

●Initial evaluation – The initial evaluation should include:

•Pregnancy history, family history, and physical examination.

•Pelvic/abdominal ultrasonography. (See 'Imaging' below.)

•Laboratory evaluation including karyotype or other rapid assessment of sex chromosome number that may include a probe for SRY and other Y and X chromosome markers using fluorescence in situ hybridization (FISH), measurement of baseline electrolytes (with repeated testing every 24 to 48 hours to evaluate for salt-wasting crisis until the diagnosis of CAH is excluded), and assessment of adrenal and gonadal function. (See 'Initial laboratory testing' below.)

●Categorization – The information from the initial evaluation can be used to categorize the infant into one of three categories, based primarily on the sex chromosome complement [5]:

•XX DSD, which is caused by virilization (table 3A)

•XY DSD, which is caused by undervirilization (table 3B)

•Sex chromosome mosaicism/chimerism (specifically, mosaicism/chimerism involving the Y chromosome)

Within each of these chromosomal categories, the results of imaging and laboratory tests for gonadal and adrenal function are used to determine the likely mechanism of the DSD and potential causal mutations (see 'Interpretation of initial findings' below). An initial recommendation for gender/sex designation is usually possible at this point.

●Second-line testing for specific disorders and associated anomalies – The selection of subsequent tests is dictated by the results of the karyotype and other results. (See 'Subsequent evaluation' below.)

PRENATAL PRESENTATION — DSD may be suspected prenatally because of atypical genital appearance on fetal ultrasound or, if prenatal determination of sex chromosomes has been done, genital appearance discordant with the sex-chromosome complement. Three-dimensional sonography can be useful to better define the findings. In one study of families referred to a DSD center because of atypical genital appearance identified on prenatal ultrasound, two-thirds also had other, non-genital anomalies, a DSD was confirmed postnatally in approximately 80 percent, and a genetic cause was diagnosed in 24 percent [13]. Genetic counseling and referral to a DSD center should be offered, with the option of prenatal genetic testing if desired by the family. (See 'Genetic testing' below.)

HISTORY — The history in a child with a DSD should include the following information:

●Prenatal exposure to androgens (eg, danazol, testosterone, synthetic progestins) or endocrine disrupters (eg, phenytoin, phenobarbital, spironolactone, finasteride). (See "Gestational hyperandrogenism" and "Risks associated with epilepsy during pregnancy and the postpartum period", section on 'Effects of ASMs on the fetus and child'.)

●Maternal virilization in pregnancy (which could indicate placental aromatase deficiency or a maternal androgen-secreting tumor). (See "Gestational hyperandrogenism".)

●Maternal family history of women who have been unable to bear children and/or have amenorrhea (which could indicate androgen insensitivity). (See "Diagnosis and treatment of disorders of the androgen receptor".)

●History of consanguinity or genetically homogeneous population (which would increase suspicion for recessive disorders, eg, congenital adrenal hyperplasia [CAH] or disorders of androgen biosynthesis). (See "Genetic counseling: Family history interpretation and risk assessment", section on 'Family history'.)

PHYSICAL EXAMINATION — The physical examination should include careful inspection and palpation of the genitalia. The labioscrotal folds and inguinal regions should be palpated for gonads and the number of urogenital openings documented. Measurements of the phallus/clitoris and anogenital ratio should be obtained; the specific measurement technique and norms are described below. (See 'Clitorophallic size' below and 'Labioscrotal fusion and the anogenital ratio' below.)

Manifestations of a DSD may include bilateral cryptorchidism (picture 2), scrotal or perineal hypospadias (picture 3), clitoromegaly (picture 5A-B), posterior labial fusion and/or other virilization of generally female-appearing genitalia (which, if severe, can present as a common urogenital sinus) (picture 6), female-appearing genitals with a palpable gonad, or hypospadias with a unilateral nonpalpable gonad (picture 4A-B). Each of these findings has a pathophysiologic basis that can provide a clue to the underlying cause of the DSD.

Virilization scales — The external genitalia score provides a method for systematically documenting genital phenotype [14]. Other scales include the Prader scale developed for XX children with congenital adrenal hyperplasia (CAH) (figure 4A-B) and the Quigley scale for XY children with androgen insensitivity syndrome (figure 5) [15]. These scales can be useful for systematically documenting genital phenotype.

Clitorophallic size

●Measurement – Clitorophallic length is measured in the nonerect state on the dorsal surface from the pubic ramus to the tip of the clitorophallus (excluding any excess foreskin or clitoral hood tissue) after stretching to the point of increased resistance. An object such as a ruler with rounded corners or a tongue depressor should be pressed down against the ramus to completely depress the suprapubic fat pad, which can conceal part of the phallic shaft. Curvature and/or ventral tethering of the clitorophallus (chordee) may interfere with the measurement, in which case, it may be necessary to estimate the length. Clitorophallic width (diameter) is measured at the midshaft.

If it is difficult to isolate the clitorophallus, palpating the spongy corporal tissue may help to distinguish the clitorophallus from surrounding tissue and allow a rough estimate of size.

●Interpretation – Standards for stretched penile length and for clitoral size from infancy through adulthood are available [5]. In a typical XY term infant, penile length is ≥2.5 cm and penile diameter measured at the middle of the shaft is ≥0.9 cm. These measurements should be adjusted for gestational age (figure 6) [16]. In XY individuals, reduced clitorophallic size can result from a DSD causing insufficient androgen action during the first trimester and is usually associated with hypospadias and incomplete fusion of the labioscrotal folds. Micropenis (microphallus) in XY individuals may also be caused by deficiencies of growth hormone and/or gonadotropins during the second and third trimesters, as can be seen in idiopathic hypogonadotropic hypogonadism/Kallmann syndrome, isolated growth hormone deficiency, and panhypopituitarism. Micropenis with accompanying hypospadias, however, is rarely secondary to growth hormone or gonadotropin deficiency.

Clitoral width in a typical XX neonate ranges from 2 to 6 mm. Clitoral length in the newborn infant may vary in different population groups, but lengths of more than 9 mm are unusual [17-19]. The clitoris may appear disproportionately more prominent in preterm infants because clitoral size is fully developed by 27 weeks of gestation but there is less fat in the labia majora than in term infants [20].

In XX individuals, frankly increased clitorophallic size is caused by inappropriate androgen action.

Gonadal location — The scrotum, labia majora, and inguinal area should be carefully palpated to identify the presence and position of the gonads.

●Interpretation – Gonads palpable below the inguinal ligament (eg, in the inguinal region or in the labioscrotal folds) (picture 7) have at least some testicular tissue [21]. This is usually associated with the presence of a Y chromosome in at least some cells (XY karyotype or a karyotype with Y chromosome mosaicism/chimerism), though, in rare cases, a palpable inguinal gonad can be seen with XX testicular or ovotesticular DSD or the mass may be part of the uterus protruding into an inguinal hernia.

All infants with atypical genital appearance and nonpalpable gonads should be promptly evaluated for CAH even before the results of the karyotype are available to avoid the complication of a potentially life-threatening adrenal crisis. CAH causing virilization of an XX child is the most common cause of atypical genitalia. Only rare forms of CAH cause undervirilization of an XY child, so a finding of one or two palpable gonads makes CAH much less likely. (See 'Initial laboratory testing' below and "Undescended testes (cryptorchidism) in children: Clinical features and evaluation", section on 'Bilaterally nonpalpable testes'.)

In an XY newborn, bilateral nonpalpable testes may occur not only with a DSD but also with congenital anorchia (testicular regression/vanishing testes syndrome) or isolated cryptorchidism, which is most commonly idiopathic but may be a manifestation of persistent müllerian duct syndrome or other causes.

Asymmetry of the gonads or other genitalia may indicate a difference between the identities of the right and left gonads, as can be seen in mixed gonadal dysgenesis (picture 4A).

Urethral opening — The genitals should be inspected for the location of the urethral opening (ideally confirmed by observing urination from the orifice) and for the presence or absence of a separate vaginal opening. However, the physical examination can sometimes be misleading; for example, a blind-ending dimple at the tip of the glans can be mistaken for the urethral meatus.

A single opening at the base of the phallus (picture 8) may be either a misplaced penile urethra (hypospadias) or a virilized urogenital sinus (eg, internal connection between the vagina and urethra), both indicating partial but incomplete androgen action. Ultimately, the physical findings can be confirmed either by cystoscopy/vaginoscopy or radiographically. (See 'Imaging' below.)

Labioscrotal fusion and the anogenital ratio — The anogenital ratio, which is independent of gestational age and body size, is the distance between the anus and posterior fourchette divided by the distance between the anus and the base of the clitoris/phallus (figure 3).

●Interpretation – A ratio of >0.5 (or >0.6) indicates posterior labial fusion due to androgen action [19,22]

Other physical features — Associated nongenital anomalies or dysmorphic features should be documented (table 4). The presence of associated nongenital anomalies usually excludes common forms of CAH but may occur in patients with P450 oxidoreductase deficiency.

As examples:

●Elevated blood pressure can be seen with 11-beta-hydroxylase deficiency and 17-hydroxylase deficiency but is not always a consistent finding in infancy.

●If gastrointestinal anomalies accompany atypical genital development, a disorder of cloacal differentiation is most likely. (See "Body stalk anomaly and cloacal exstrophy: Prenatal diagnosis and management".)

●Infants with Smith-Lemli-Opitz syndrome (a disorder of cholesterol biosynthesis caused by deficiency of 7-dehydrocholesterol reductase, encoded by the DHCR7 gene) may have a variety of phenotypic abnormalities in addition to atypical genital appearance (picture 9). These include microcephaly, micrognathia, low-set and posteriorly rotated ears, and syndactyly and/or polydactyly [23]. (See "Causes of differences of sex development", section on 'Smith-Lemli-Opitz syndrome' and "Causes of primary adrenal insufficiency in children", section on 'Defects in cholesterol biochemistry'.)

●Individuals with P450 oxidoreductase deficiency, a rare form of CAH, may have craniofacial and limb abnormalities (also known as Antley-Bixler syndrome). (See "Uncommon congenital adrenal hyperplasias", section on 'P450 oxidoreductase deficiency (apparent combined CYP17A1 and CYP21A2 deficiency)'.)

INITIAL LABORATORY TESTING — The initial laboratory evaluation of the infant with a DSD includes (table 2):

●Expedited evaluation of sex chromosomes – Using karyotype or fluorescence in situ hybridization (FISH) for SRY and other markers of the X and Y chromosomes, depending on which method will return results most quickly.

●Assessment of gonadal function at the appropriate time after birth – Follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone, dihydrotestosterone, and anti-müllerian hormone (AMH).

●Measurement of adrenal steroids – 17-hydroxyprogesterone (17-OHP). Additional tests for uncommon causes of congenital adrenal hyperplasia (CAH; 17-hydroxypregnenolone, cortisol, and 11-deoxycortisol) are sometimes done at this point but may be deferred to minimize blood loss. (See '17-hydroxyprogesterone' below and 'Other assessment of adrenal function' below.)

●Measurement of baseline electrolytes – In patients with confirmed or possible CAH. Electrolyte measures should be repeated every 24 to 48 hours to monitor for impending adrenal crisis, until the diagnosis of CAH is excluded. (See 'Electrolytes' below.)

Sex chromosome analysis — Analysis of the sex chromosomes is a critical element of diagnosis and should be performed as soon as possible in any infant with atypical genital appearance. If expedited assessment of the sex chromosomes is not available locally, transfer to a facility with access to a rapid turnaround time should be strongly considered.

●Method – Because of the possibility of mosaicism/chimerism, it is suggested that at least 50 cells be examined. In some centers, methods such as FISH with X and Y chromosome probes may offer a faster way to determine the sex chromosome complement if the karyotype result will be slow to return. Both karyotype and FISH can provide additional useful information, such as the presence of deletions, duplications, and translocations with karyotype, and the presence/absence of the SRY gene with FISH. (See 'Genetic testing' below.)

If the sex chromosome number was evaluated as part of prenatal screening, the results should be confirmed on a sample from the infant or from cord blood. Although an assessment of sex chromosome number performed prenatally is generally reliable, inaccurate results are possible:

•Cell-free (free fetal) DNA tests assess the chromosomal complement of the placenta, which may or may not be the same as that of the fetus, particularly if there is mosaicism. Furthermore, cell-free DNA tests were developed for the detection of autosomal trisomies, not for detection of sex chromosome anomalies.

•Chorionic villus sampling similarly assesses placental chromosomes.

•Amniocentesis assesses the fetal chromosomes and is the most reliable of the available prenatal tests but may still give misleading results in rare situations (eg, fetal twin demise).

It is possible that mosaicism/chimerism affects the gonads but does not extend to the hematopoietic lineage. In these rare situations, a karyotype of the gonadal tissue obtained when surgery is being done for other reasons or analysis of buccal swabs or skin fibroblasts may be helpful in identifying mosaicism/chimerism. Conversely, case reports describe an inaccurate diagnosis of 46,XY DSD due to hematologic chimerism in sex-discordant twins due to twin-twin transfusion [24].

●Interpretation – The results of the karyotype permit classification of the infant into one of three diagnostic categories that guide further evaluation [5]:

•XX DSD

•XY DSD

•Sex chromosome DSD – With a normal or aberrant Y chromosome present in some but not all cells

17-hydroxyprogesterone — 17-OHP is measured to evaluate for 21-hydroxylase deficiency, which is the most common type of CAH. Undiagnosed, and hence untreated, CAH due to 21-hydroxylase deficiency can lead to life-threatening adrenal insufficiency within the first weeks of life. (See "Clinical manifestations and diagnosis of classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency in infants and children".)

●Indications – 17-OHP should be measured in all infants with bilateral nonpalpable gonads presenting with atypical genital appearance. This test is also appropriate for term infants with bilateral nonpalpable gonads and typical male genital appearance because, very rarely, XX infants with 21-hydroxylase deficiency have sufficiently marked virilization to result in complete virilization of the external genitalia. Measurement of 17-OHP should be done after 48 hours of life to avoid the birth-associated surge in adrenal hormones.

Neonatal screening for 21-hydroxylase deficiency (through measurement of 17-OHP) is routinely performed in all of the United States and in many other countries. However, because of the risk of adrenal crisis, any infant presenting with atypical genitalia should have a rapid and extensive evaluation for CAH, as described above, without waiting for the results of the newborn screen. Moreover, there may be false-negative results for neonatal screening for CAH [25]. (See "Clinical manifestations and diagnosis of classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency in infants and children".)

●Interpretation – Nomograms are available for the interpretation of unstimulated and stimulated 17-OHP [26]. A markedly elevated 17-OHP is diagnostic of 21-hydroxylase deficiency. Moderate elevations of 17-OHP may also indicate 21-hydroxylase deficiency or other types of CAH but may also occur due to prematurity and/or stress (particularly if measured within the first 48 hours of life). If the result of the 17-OHP test is not definitive, adrenocorticotropic hormone (ACTH) stimulation testing may be required to establish a definitive diagnosis.

Electrolytes — Electrolytes are measured to monitor for the possibility of a salt-wasting adrenal crisis, a potentially life-threatening complication of CAH. Salt-wasting adrenal crisis does not occur in the first few days of life, with extremely rare exceptions. This affords some time to establish the diagnosis of CAH, but initial normal electrolytes also do not exclude the possibility of adrenal insufficiency.

●Indications – In any infant for whom CAH is considered possible, serum electrolytes should be measured at the time of presentation with atypical genitalia and then repeated every 24 to 48 hours until CAH is excluded.

●Interpretation – Salt wasting is suggested by the findings of hyponatremia, hyperkalemia, and nongap metabolic acidosis, the hallmarks of adrenal insufficiency with mineralocorticoid deficiency. However, elevated serum potassium measurements may also be caused by hemolysis, which is a common problem with blood collection in infants.

Other assessment of adrenal function — Other steroid hormones and precursors should be measured to evaluate for uncommon types of CAH, which can cause atypical genitalia in either XX or XY infants. These are rare disorders but can be important to diagnose early since some are associated with salt wasting and adrenal crisis.

●Tests:

•Testing for less common types of CAH involves measuring dehydroepiandrosterone (DHEA), 17-hydroxypregnenolone (not to be confused with 17-hydroxyprogesterone [17-OHP]), 11-deoxycortisol, cortisol, and potentially other intermediates.

•Measurement of ACTH can be useful in demonstrating primary adrenocortical insufficiency due to CAH or adrenal dysgenesis related to defects in the NR5A1 (SF1) gene.

●Interpretation – Abnormal results suggest a type of adrenal dysfunction (table 5):

•Elevated 11-deoxycortisol concentration suggests 11-beta-hydroxylase deficiency, which can cause virilization of XX infants.

•Elevated 17-hydroxypregnenolone and an elevated ratio of 17-hydroxypregnenolone to cortisol suggest 3-beta-hydroxysteroid dehydrogenase deficiency, which can cause virilization of XX infants and undervirilization of XY infants.

•Elevated ACTH suggests primary adrenocortical insufficiency, which may be associated with CAH or with homozygous NR5A1 gene mutations, a cause of both adrenal and gonadal dysgenesis. ACTH can be physiologically elevated in stressed or ill infants.

Testosterone and other measures of gonadal function — Unstimulated values of testosterone, dihydrotestosterone, LH, FSH, and AMH should be measured in all infants with atypical genitalia; the tests and interpretation are outlined in the table (table 2). The method and timing of measurement are critical to ensure valid results:

●Method – To ensure accuracy, testosterone assays in infants must be performed using gas or liquid chromatography-mass spectroscopy (LC/MS) rather than standard immunoassay. Alternatively, adding an extraction step prior to immunoassay may produce accurate results. Cross-reacting steroids from the maternal circulation and the fetal adrenal cortex may falsely elevate the results of immunoassays if performed without extraction and careful preparation. This fact has greatly confused the literature.

●Timing – The testosterone assay should ideally be performed after one week of age and before six months of age. Samples taken during the first several days of life may not be a valid reflection of testicular function, because the reproductive endocrine axis activity is suppressed at birth (although it is highly active during fetal life). Levels of testosterone begin to rise by approximately one week of age in full-term infants, marking the start of the "mini-puberty" of infancy. In boys, serum testosterone usually peaks between one to three months and male mini-puberty ends by approximately six months of age. In girls, mini-puberty ends by around two years of age [27-29]. Premature infants may take longer to enter the "mini-puberty" phase.

●Associated tests – LH, FSH, and AMH are interpreted as follows:

•LH is the stimulus for testosterone production by the Leydig cells of the testis (in individuals with Leydig cells) and serves as an index of hypothalamic-pituitary function (regardless of karyotype or gonadal development).

An LH concentration above 0.3 mIU/mL demonstrates that the infant has entered "mini-puberty" and is therefore capable of stimulating production of testosterone if Leydig cells are present [30-32]. Low LH can be seen in children who are not in mini-puberty, children with hypogonadotropic hypogonadism (which could be due to a pathologic condition or due to a normal physiologic response to stress), and, for reasons that are unclear, some children with complete androgen insensitivity syndrome [33]. If LH is low and the infant is younger than 14 days, testing should be repeated later, when the child is more likely to be in mini-puberty. Alternatively, human chorionic gonadotropin (hCG)-stimulation testing can be performed to assess testosterone and dihydrotestosterone production. (See 'Human chorionic gonadotropin stimulation test' below.)

An elevated LH during the mini-puberty of infancy suggests insufficient negative feedback from sex steroids, which can be seen with gonadal dysgenesis; defects in testosterone, dihydrotestosterone, and/or estradiol synthesis; and androgen insensitivity [27].

•In individuals with testes that contain functional Sertoli cells, AMH is produced in response to FSH.

Unlike most other gonadal hormones, AMH is still produced after mini-puberty ends, and there are clear differences in circulating AMH between boys (>30 ng/mL) and girls (<10 ng/mL) from infancy through childhood until the onset of puberty. After pubertal onset, this sexual dimorphism disappears as testosterone begins to suppress AMH production in boys and follicular development leads to increases in AMH production in girls.

In a virilized XX infant or child, an AMH level above the female range indicates the presence of Sertoli cells and, in turn, the presence of testicular tissue and therefore suggests an XX testicular or ovotesticular DSD. If AMH is below the typical male range in an XY individual with atypical genital appearance, this suggests a global defect in testicular function, ie, gonadal dysgenesis affecting both Leydig and Sertoli cells. If AMH is within the typical male range in an XY individual with undervirilization, this suggests an isolated defect in androgen synthesis or action.

In conjunction with the karyotype, these tests help to determine whether testicular tissue is present, the source of androgens in a virilized XX infant, and the level of the defect in androgen production or action in an undervirilized XY infant. (See 'Interpretation of initial findings' below.)

Imaging — Ultrasonography of the abdomen and pelvis, and occasionally magnetic resonance imaging or genitoscopy, is important to determine the presence of gonads, a uterus, and/or a vagina.

●Interpretation – Taken at face value, the absence of a uterus suggests sufficient AMH production by Sertoli cells to cause müllerian regression. In a patient with impalpable gonads, inability to visualize gonads on ultrasound suggests gonadal dysgenesis. However, the ability to detect a uterus and/or abdominal gonads will vary by facility and false-negative and false-positive results are possible, so concurrent measurement of AMH is recommended. It is sometimes but not always possible to determine whether gonads appear more testicular or ovarian. Adrenal hyperplasia associated with CAH can sometimes be appreciated but is not a sufficiently reliable finding to establish or exclude the diagnosis.

Interpretation of initial findings — The results from the initial evaluation of the child with atypical genital development should reveal the sex chromosome complement, allow the identity of the gonads to be inferred, and suggest potential underlying causes.

For individuals with XX or XY sex chromosomes, the initial tests can be used to narrow the differential diagnosis and to select further testing, as outlined in the table (table 2):

●XX – In XX individuals, the fundamental question is whether the virilization is due to androgens from the adrenal glands (due to CAH [most common]), the gonads (due to testicular or ovotesticular DSD [rare]), or another source (table 3A):

•Adrenal androgen excess (CAH) can generally be diagnosed on the basis of abnormalities in adrenal precursors.

•Gonadal overproduction of androgens is most often due to XX testicular or ovotesticular DSD. These conditions are caused by defects in gonadal (ovarian) development and can usually be recognized by the finding of testosterone and AMH above the normal female range, often accompanied by absence or hypoplasia of müllerian structures due to AMH action. A rare cause of gonadal overproduction of androgens is aromatase deficiency.

•Gestational hyperandrogenism (which should be apparent from a history of maternal virilization during pregnancy).

●XY – In XY individuals, the fundamental question is why there was insufficient action of testosterone and/or dihydrotestosterone. The causes can be broadly divided into the following categories (table 3B):

•Conditions that affect testicular function globally – ie, gonadal dysgenesis. In these disorders, there may be decreased production of not only testosterone but also AMH, which can in turn lead to retention of müllerian structures.

•Conditions that affect dihydrotestosterone synthesis or action specifically – ie, disorders of testosterone synthesis, 5-alpha-reductase deficiency, and androgen insensitivity. In these disorders, Sertoli cell function is intact and production of AMH results in absence of müllerian structures and serum AMH in the typical male range; furthermore, production of inhibin B results in partial feedback inhibition of FSH, such that LH is typically elevated more than FSH. The serum testosterone and dihydrotestosterone concentrations (random or stimulated by hCG) can then be used to determine whether there is a defect in dihydrotestosterone synthesis or androgen insensitivity is present, though characteristic biochemical abnormalities are not always present and genetic testing is sometimes needed to establish a definitive diagnosis.

●Sex chromosome DSD – A finding of mosaicism/chimerism resulting in the presence of the Y chromosome in some but not all cells (eg, a 45,X/46,XY karyotype) is sufficient to explain atypical genital development. This can result in a broad range of reproductive phenotypes, including normal or dysgenetic testes or ovaries, streak (severely dysgenetic) gonads, ovotestes, or mixed gonadal dysgenesis (gonads that develop differently from each other, with at least one being dysgenetic). (See "Causes of differences of sex development", section on 'Sex chromosome differences of sex development'.)

With this information, recommendations for sex/gender designation can usually be made (see "Management of the infant with atypical genital appearance (difference of sex development)"). However, some conditions (such as 17-beta-hydroxysteroid dehydrogenase [17-beta-HSD] deficiency) may not be apparent on initial testing and may take longer to diagnose and, in rare circumstances, the initial recommendation may need to be revised later.

SUBSEQUENT EVALUATION — With the initial evaluation complete, subsequent laboratory testing is generally performed to:

●Confirm a suspected diagnosis (eg, sequencing of the AR gene to confirm suspected androgen insensitivity)

●Identify specific causes within a general category of DSD (eg, if gonadal dysgenesis is diagnosed, sequencing the NR5A1 (SF1) gene to identify mutations)

●Determine the identity and functionality of the gonads

●Screen for associated nongenital anomalies (eg, echocardiography to screen for cardiac defects in individuals with a 45,X karyotype in some cells)

Other steroid precursors — In many cases, the initial evaluation will definitively identify a cause of the DSD, such as congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency or Y chromosome mosaicism, and no further diagnostic testing is needed.

If the diagnosis was not established with initial testing, then measurement of other adrenal and/or gonadal steroid precursors (if not already done) should be performed to identify rare forms of CAH, which can cause atypical genitalia in either XX or XY infants (table 5). Tests include 11-deoxycortisol, 17-hydroxypregnenolone, and cortisol; the interpretation is outlined above. (See 'Other assessment of adrenal function' above.)

Other causes of CAH are exceedingly rare, and diagnoses of these conditions requires measurement of other adrenal precursors, often following an adrenocorticotropic hormone (ACTH) stimulation test.

In XY infants with testicular tissue, measurement of the testosterone:androstenedione ratio may help to identify deficiency of 17-beta-hydroxysteroid dehydrogenase (17-beta-HSD), in which the ratio is typically <0.8. The diagnostic accuracy may be enhanced by human chorionic gonadotropin (hCG) stimulation testing, but some cases can be diagnosed only through genetic testing. (See 'Human chorionic gonadotropin stimulation test' below and "Causes of differences of sex development", section on '17-beta-hydroxysteroid dehydrogenase type 3 deficiency'.)

Stimulation testing — For selected patients, stimulation testing may be needed to establish or confirm a diagnosis and/or to identify associated issues.

Adrenocorticotropic hormone stimulation test — An ACTH stimulation test is used to identify defects in adrenal hormone synthesis and/or to establish or exclude adrenocortical insufficiency in selected patients. It is not needed for the majority of XX infants with classical CAH (21-hydroxylase deficiency), because their basal levels of 17-hydroxyprogesterone (17-OHP) are usually sufficiently elevated to establish the diagnosis. Similarly, basal levels of other adrenal hormone precursors are often (but not always) sufficient to establish a diagnosis.

●Indications – An ACTH stimulation test should be performed in:

•XX infants with borderline elevations in 17-OHP

•XX infants with suspected but unconfirmed 3-beta-hydroxysteroid dehydrogenase deficiency or 11-beta-hydroxylase deficiency

•XY infants with suspected but unconfirmed 17-alpha hydroxylase or 3-beta-hydroxysteroid dehydrogenase deficiency

•Infants with other adrenal steroidogenic defects, if initial laboratory studies with baseline cortisol levels have not clearly demonstrated a capacity to produce cortisol with stress

The main goals of this test are to confirm the biochemical diagnosis if no genetic diagnosis is available and to determine if there is associated adrenal insufficiency.

●Technique – We perform the ACTH stimulation test in the following manner:

•Measure ACTH, cortisol, pregnenolone, progesterone, 17-alpha-hydroxypregnenolone, 17-alpha-hydroxyprogesterone, 11-deoxycortisol, dehydroepiandrosterone (DHEA), and androstenedione.

•Administer ACTH (a supraphysiologic dose, such as 62.5 mcg of synthetic ACTH).

•Sixty minutes after administration of ACTH, measure cortisol, pregnenolone, progesterone, 17-alpha-hydroxypregnenolone, 17-alpha-hydroxyprogesterone, 11-deoxycortisol, DHEA, and androstenedione. These assays may be obtained as a panel in some laboratories.

●Interpretation – Test results should be interpreted using the age-specific normal values provided by the laboratory performing the testing.

Failure of any steroidogenic response suggests a condition affecting adrenal development (such as a mutation in the NR5A1 [SF1] gene) or a condition affecting an early step in adrenal steroidogenesis (due to mutations in the STAR or CYP11A1 [P450SCC] genes) (table 5). Abnormal levels and/or ratios of steroidogenic precursors may point to a specific type of CAH. (See "Uncommon congenital adrenal hyperplasias", section on 'Lipoid congenital adrenal hyperplasia' and "Causes of primary adrenal insufficiency in children", section on 'Congenital adrenal hyperplasia'.)

Human chorionic gonadotropin stimulation test

●Indications – In a patient who has testicular tissue on ultrasound and/or normal concentrations of anti-müllerian hormone (AMH), the hCG stimulation test can be performed to try to distinguish between disorders of abnormal androgen synthesis and those with abnormal androgen sensitivity [34]. In the past, hCG stimulation tests were used to determine the presence of testicular tissue (eg, to evaluate for testicular regression syndrome), but this has largely been supplanted by measurement of AMH [35]. If blood samples for testosterone and dihydrotestosterone can be obtained during the mini-puberty of infancy (approximately age two weeks through six months), hCG testing is not usually necessary. Furthermore, as genetic diagnoses become more common, the hCG stimulation test may be used even less.

●Technique – There are numerous protocols for hCG stimulation testing; most protocols use two to three doses of hCG given daily or every other day and laboratory measurement at baseline and one to three days after the last dose of hCG. An example of a protocol is:

•Basal luteinizing hormone (LH), follicle-stimulating hormone (FSH), androstenedione, testosterone, and dihydrotestosterone are measured

•Recombinant hCG (1500 units/m² subcutaneously) is administered on day 1 and repeated on day 3

•Beta-hCG, androstenedione, testosterone, and dihydrotestosterone are measured on days 3 and 6; measurement of beta-hCG confirms that the medication has been administered

●Interpretation – The response of normal Leydig cells to hCG is usually quite robust. However, widely varying results are reported in the literature depending upon subject age and puberty status and the technique of the hCG test (including dose amount and number and timing of stimulated blood sample after hCG administration). A ratio of testosterone:dihydrotestosterone (when expressed in the same units) after the administration of hCG >10:1 suggests 5-alpha-reductase 2 deficiency [34]. A ratio of testosterone:androstenedione following hCG stimulation <0.8 [36] suggests 17-beta-HSD3 deficiency, but some individuals with 17-beta-HSD3 deficiency will have a normal hormonal profile (due to conversion of androstenedione to testosterone in the periphery by isoforms of 17-beta-HSD encoded by other genes) (table 3B). (See "Causes of differences of sex development", section on 'Reduced androgen synthesis'.)

Identifying ovarian tissue — In rare cases (eg, when an ovotesticular DSD is suspected based on the appearance of the gonads on ultrasonography), there may be a question of whether ovarian tissue is present.

Measurement of inhibin A may be useful for demonstrating the presence of ovarian tissue in infants, but this is based on limited evidence and is rarely used. Inhibin A is secreted by the ovary but not the testis, and circulating inhibin A is detectable in female infants in the first two months of life [37].

Administration of human menopausal gonadotropins to stimulate ovarian function also might be used as a test to identify ovarian tissue, measuring either an increase in estrogen levels or ultrasound evidence of follicular development [38,39]. One study has shown that FSH-stimulated inhibin A levels can identify the presence of ovarian tissue in children [37]. These approaches could be used in infants who are older than two or three weeks of age. However, evidence for the use of inhibin A to establish the presence of ovarian tissue in infants is limited.

Genetic testing — Many different genes have been associated with DSDs, as outlined in the tables (table 3A-B).

●Candidate gene testing – Genetic testing for specific candidate genes may be prompted by the clinical picture. For example:

•For patients with clinical features and a family history suggesting androgen insensitivity, test for the AR gene, which encodes the androgen receptor. (See "Causes of differences of sex development", section on 'Androgen insensitivity'.)

•For patients with suspected 46,XX testicular or ovotesticular DSD, test for the SRY gene using fluorescence in situ hybridization (FISH) because the presence of SRY accounts for a significant proportion of such cases [40]. Evaluation of SRY can also be considered in individuals with XY gonadal dysgenesis, either by FISH to detect deletions or by sequencing to detect point mutations. (See "Causes of differences of sex development", section on 'Gonadal overproduction of androgens' and "Causes of differences of sex development", section on 'XY gonadal dysgenesis'.)

•For patients with suspected XY gonadal dysgenesis or XX testicular or ovotesticular DSD, test for mutations in the NR5A1 (SF1) gene, which are found in 10 to 15 percent of cases. (See "Causes of differences of sex development", section on 'Gonadal overproduction of androgens' and "Causes of differences of sex development", section on 'XY gonadal dysgenesis'.)

•For patients with suspected 17-beta-HSD deficiency (eg, XY individuals with low-normal serum testosterone concentrations and elevated androstenedione), test for mutations in the HSD17B3 gene. This disorder may not produce classical hormonal changes and is sometimes discovered only on the basis of genetic testing of the HSD17B3 gene. (See "Causes of differences of sex development", section on '17-beta-hydroxysteroid dehydrogenase type 3 deficiency'.)

●Multigene sequencing – Because many genetic causes of DSD are rare, gene-by-gene testing can be expensive and of low yield. DSD gene panels and comprehensive sequencing methods such as whole-exome and whole-genome sequencing are becoming increasingly available and cost-effective. Such approaches are able to identify a genetic cause of DSD in 20 to 50 percent of cases that are not due to CAH [41-50]. (See "Tools for genetics and genomics: Cytogenetics and molecular genetics", section on 'Array comparative genomic hybridization' and "Next-generation DNA sequencing (NGS): Principles and clinical applications".)

●Karyotype and/or chromosomal microarray analysis (CMA) – Karyotype, if not previously done to determine the sex chromosome complement, can detect large deletions and duplications as well as chromosomal translocations. CMA (also known as array-based comparative genomic hybridization [aCGH]) detects duplications and deletions that may be too small to be identified on karyotype. Some DSDs have been found to be due to such deletions or duplications (eg, SOX3 or SOX9 gene duplications in 46,XX testicular DSD and NR0B1/DAX1 gene duplications in 46,XY testicular dysgenesis) [51-53]. Studies suggest that CMA can identify a genetic cause in 10 to 30 percent of DSD cases that are not due to CAH [51-53].

●Our approach – In our practice, if the clinical and biochemical picture does not suggest a specific molecular diagnosis, we perform multigene sequencing with a DSD gene panel or whole-exome sequencing (if available). We perform multigene sequencing rather than CMA because the yield appears to be higher; we reserve CMA for cases in which screening for coding mutations is negative. However, multigene sequencing is often not covered by insurance and may be difficult to obtain in clinical settings. Therefore, test availability and insurance coverage may influence the order of testing. If testing single genes one by one is necessary, the most common genetic variants for XX DSD are the presence of SRY and coding variants in NR5A1. For XY DSD, the most common genetic variants are coding variants in AR and NR5A1.

We anticipate that, at some point, genetic screening will be sufficiently rapid and inexpensive that it may become first-line in the diagnostic approach to DSD and replace some of the biochemical studies discussed above, although many of the biochemical tests will remain important for defining the severity of the condition and for management.

Second-line imaging — Retrograde urethrogram or genitogram may be helpful in defining internal anatomy (eg, the presence of a urogenital sinus), but this information is primarily used to guide surgical management and thus does not typically need to be done in the neonatal period. Furthermore, most surgeons find direct visualization by cystoscopy/vaginoscopy to be the best method of assessing the urethral and vaginal anatomy. In some complicated cases (particularly those infants with elements of both male and female gonads/internal reproductive structures), laparoscopic visualization with gonadal biopsy may be required to completely inventory the reproductive structures.

Individuals with sex chromosome mosaicism and a 45,X karyotype in some cells are at risk for anomalies associated with Turner syndrome and require echocardiography, renal ultrasound, and hearing screening to screen for cardiac defects, anatomic kidney anomalies, and hearing impairment, respectively. (See "Clinical manifestations and diagnosis of Turner syndrome".)

SOCIETY GUIDELINE LINKS — Links to society and government-sponsored guidelines from selected countries and regions around the world are provided separately. (See "Society guideline links: Classic and nonclassic congenital adrenal hyperplasia due to 21-hydroxylase deficiency" and "Society guideline links: Differences of sex development".)

SUMMARY AND RECOMMENDATIONS

●Terminology – Individuals with atypical genital development are classified as having a difference (or disorder) of sex development (DSD). Some DSDs present with an atypical genital appearance that does not permit gender designation at birth; others present with genital appearance that is discordant with genetic sex. (See 'Terminology' above.)

●Guiding principles – The evaluation of infants with atypical genitalia should be undertaken as soon as possible. This is because congenital adrenal hyperplasia (CAH), the most common cause of DSD, can be life-threatening. In addition, a DSD is perceived as distressing by most families and calls for immediate sensitive and professional counseling and psychosocial support. (See 'Guiding principles' above.)

●Initial evaluation

•Physical examination – Key clinical features of infants with a DSD can include bilaterally nonpalpable testes (picture 2), microphallus, scrotal or perineal hypospadias (picture 3), clitoromegaly (picture 5A-B), posterior labial fusion (picture 6), or palpable gonad(s) in the labioscrotal folds (picture 7). (See 'Physical examination' above.)

•Laboratory tests – Initial laboratory testing should include:

-Expedited karyotype or other method to rapidly assess the sex chromosome complement.

-17-hydroxyprogesterone (17-OHP) for infants without palpable gonads, as a screen for CAH due to 21-hydroxylase deficiency.

-Serum electrolytes, measured every 24 to 48 hours until CAH is excluded, to monitor for the possibility of salt wasting due to adrenal insufficiency, which can be life-threatening.

-Because salt wasting is also associated with some less common types of CAH, there should be a low threshold for evaluating for these disorders by measuring dehydroepiandrosterone (DHEA), 17-hydroxypregnenolone, 11-deoxycortisol, cortisol, and adrenocorticotropic hormone (ACTH) (table 5), especially in any infant with evidence of salt loss. (See 'Initial laboratory testing' above.)

•Imaging – Pelvic and abdominal ultrasonography should be performed to determine whether gonads, uterus, and vagina are present. (See 'Imaging' above.)

●Interpretation and initial categorization of DSD – The results of the examination and initial laboratory testing should permit classification of the infant into one of three broad categories: XX DSD, XY DSD, or sex chromosome DSD. The infant can then be further evaluated to determine the nature of the underlying disorder within each of these categories, as outlined in the table (table 2).

•In XX infants, the most common cause of atypical genitalia is CAH due to 21-hydroxylase deficiency; the differential diagnosis also includes less common causes of CAH, gestational hyperandrogenism, testicular DSD, ovotesticular DSD, and aromatase deficiency (table 3A).

•In XY infants, the differential diagnosis includes defects in testicular development, defects in testosterone/dihydrotestosterone synthesis, and androgen insensitivity. Defects in testosterone synthesis can be seen in uncommon forms of CAH, most of which have a risk for adrenal crisis (table 5).

In addition to measurements of 17-hydroxypregnenolone and cortisol, the evaluation should include measurement of serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), anti-müllerian hormone (AMH), testosterone, dihydrotestosterone, and androstenedione (table 3B). Patients with an XY DSD without a diagnosis after laboratory evaluation should have a genetic evaluation, which can identify many causes of an XY DSD that do not result in obvious biochemical anomalies. (See 'Genetic testing' above.)

•More details on specific disorders are discussed separately. (See "Causes of differences of sex development".)

●Genetic testing – If the cause of the DSD remains unclear after the workup outlined above, the highest-yield next step is multigene sequencing (with a DSD gene panel or whole-exome sequencing), if available. (See 'Genetic testing' above.)

●Management – The management of these infants requires a multidisciplinary team including specialists from pediatric endocrinology, genetics, and pediatric surgery/urology as well as psychology, psychiatry, or social work. (See "Management of the infant with atypical genital appearance (difference of sex development)".)

ACKNOWLEDGMENTS — The UpToDate editorial staff acknowledges Christopher P Houk, MD, who contributed to earlier versions of this topic review. The authors and UpToDate editorial staff also acknowledge Robert Rosenfield, MD, for contributing his wisdom to the discussion of steroid measurements in infants.