Diagnosis of Ambiguous Genitalia in the Newborn
When a genital defect is discovered in the neonatal period, complementary clinical, hormonal, genetic, molecular and radiographic investigations are needed to first diagnose the intersex state and then to determine the etiology and orient the therapeutic approach.
Thorough physical examination is the key to diagnosis. Methodical general inspection will establish whether the genital defect is a sign of a malformation syndrome. Careful genital inspection will define the extent of the ambiguity by determining the presence, number, size, symmetry and position of gonads. In some cases, the sexual ambiguity is evident. For example, the genital tubercle may show development that is midway between that of a penis and a clitoris (the length and diameter, however, should be evaluated); the genital folds may be completely fused, with a bifid scrotum; the penis may appear abnormally bent and buried or sunken inward; or the urethral orifice may open toward the inner side of the penis, indicating hypospadias. Other signs include posterior fusion of the labia majora and a single perineal orifice at the base of the genital tubercle, between the genital folds, signaling the existence of a urogenital sinus.
Quite often, however, the genital malformation is less obvious. When the malformation is not easy to determine, the clinician’s attention should be drawn to the following: an association of cryptorchidism and hypospadias, bilateral cryptorchidism, true clitoridal hypertrophy of the gland and not of the hood, and an oblong mass in the inguinal position in a newborn with female phenotype.
Precise measurement of the penis should be made. The mean stretched penile length in the normal full term newborn male is 3.5 cm (±0.5 cm).
Once the examination has been completed, the clinician should be ready to classify the degree of ambiguity of the external genitalia (genital tubercle and folds and the urogenital sinus). Prader’s five stages may be used or, as is now more often the case, Quigley’s 6–7 stages, which offer the advantage of greater descriptive detail.
Grade 1: Individuals with normal male external genitalia such as infertile males with azoospermia and hormonal features of androgen resistance, those with reduced virilization at puberty (so-called ‘minimal’ androgen resistance).
Grade 2: Individuals who have a univocally male phenotype, but who have mildly defective fetal masculinization, manifested by defects such as isolated hypospadias and/or micropenis.
Grade 3: Individuals with predominantly male phenotype but with more severely defective masculinization in utero, as evidenced by perineal hypospadias, small penis, and cryptorchidism and/or bifid scrotum.
Grade 4: Individuals with ambiguous phenotype and severely limited masculinization evidenced by a phallic structure that is intermediate between a clitoris and a penis, generally accompanied by a urogenital sinus with perineal orifice and labioscrotal folds.
Grade 5: Individuals with essentially female phenotype (i.e., minimal fetal androgen action), including separate urethral and vaginal orifices, with minimal androgenization evidenced by mild clitoromegaly or a small degree of posterior labial fusion.
Grade 6: Individuals with a normal female genital phenotype (i.e., no fetal androgen action) who develop androgen-dependent pubic and/or axillary hair at puberty.
Careful palpation to locate gonads below the genital folds or in the inguinal region provides the first element for diagnostic orientation. If the gonads are absent, a diagnosis of female pseudohermaphroditism seems advisable.
If a gonad or gonads are palpated, a diagnosis of male pseudohermaphroditism is indicated. It should be borne in mind, however, that cases presenting with very similar clinical expression may be quite different diagnostically as, for example, a masculinized female newborn with congenital adrenal hyperplasia (CAH) and an undervirilized male newborn.
A careful family history should be obtained from the parents. Detailed information on ambiguous genitalia in other siblings or family members, history of neonatal death, and consanguinity should all be sought. Cases of amenorrhea or infertility in the family will be important elements, and careful attention should be paid to clues of maternal ingestion of drugs or exposure to chemical environmental disruptors during pregnancy. Questions about ‘salt losing’ in the family should also be raised.
A differential diagnosis should be formulated by the end of the clinical examination (
fig. 1). A certain number of complementary investigations will then need to be carried out, although perhaps not all need be done in a systematic fashion. Testing for salt losing, however, is mandatory.
Cytogenetic and Molecular Investigations
Buccal smears reveal the presence of Barr bodies (chromatin-positive) equal to the number of X-1. In practice, this screening test is easy and rapid but insufficiently reliable – and thus it is being increasingly replaced by molecular studies of the X or Y chromosome. Karyotyping is systematic, but results are often only available several days later. Because of the urgent need for sex assignment – it is intolerable for parents to wait several weeks to know if their baby is a boy or girl – many hospitals are now also performing PCR analysis of the SRY gene on the Y chromosome, because the results are available within 1 day.
Hormonal investigations should be based on the clinical and cytogenetic orientation; although most are generally easy to perform, they can at times be quite complex. Certain hormones should be measured on an urgent basis – immediately. Other measurements should be made as quickly as possible, and still others can be made within a few days.
Substantial elevation in 17-hydroxyprogesterone (17-OHP) (and plasma testosterone, which is of lower amplitude) will confirm the diagnosis of CAH, which is due in most cases to a deficiency in 21-hydroxylase.
Basal plasma testosterone levels evaluate the presence of functional Leydig cells. Testicular stimulation with hCG (1,000 U/day for 3 days or
1,500 U every 2 days for 2 weeks) is required to determine the functional value of testicular tissue. An insufficient response (<3 ng/ml) suggests a final diagnosis of gonadal dysgenesis. This same test is also needed to show evidence of an inborn error of testosterone biosynthesis by an augmentation in the precursors (17-OHP, dehydroepiandrosterone, Δ4-androstenedione), which contrasts with the absence of variation in the plasma testosterone.
In all cases of undermasculinization of the external genitalia that is associated with an often elevated secretion of testosterone, peripheral androgen receptivity must be investigated. Depending on the group, this is accomplished either by evaluation of the clinical response of the genital tubercle to exogenous testosterone or by measurement of the concentration of receptor sites in the external genitalia.
There is no consensus regarding the dosage, method of administration, timing or duration of therapeutic trials in the newborn with micropenis and ambiguous genitalia. To test for a clinical response to testosterone, an intramuscular injection of 25 mg every 4 weeks for 3 months or 100 mg every 4 weeks for 3 months should bring about an augmentation in length, diameter, and the cavernous bodies of the genital tubercle. An augmentation in phallic length <35mm is insufficient. Failure to respond implies end-organ resistance to the action of androgens and introduces uncertainty concerning the appropriate sex of rearing.
For the evaluation of androgen receptor concentration, the number of receptors and their affinity for testosterone are measured on cultured genital skin fibroblasts. Concentrations <300 fmol/mg of DNA suggest partial androgen insensitivity.
Exploration of the genitourinary axis is principally carried out by ultrasound and genitography. In certain cases, ultrasound confirms the presence of a uterus and ovaries. Genitography can accurately detect the level of implantation of the vaginal cavity on the urethra, an essential consideration in the choice of therapeutic strategy. Three cases can be found: the vagina opens the length of the vertical urethra, forming a long urogenital sinus; the vaginal opening is in the subvesicle position; or the vagina is located at the junction of the horizontal and vertical portion of the urethra.
Revision date: July 4, 2011
Last revised: by David A. Scott, M.D.