Small supernumerary marker chromosomes (sSMCs) have been defined as structurally abnormal chromosomes that cannot be identified or characterized unambiguously by conventional banding cytogenetics alone, and are (in general) equal in size or smaller than chromosome 20 of the same metaphase spread. As they are too small for their chromosomal origin to be considered by traditional banding techniques, molecular cytogenetic techniques (including array-based comparative genomic hybridization) are required for their characterization (1). sSMCs are found in approximately 4.4/100,000 newborns (2) and they are found in three different shapes: ring, inverted duplicated and centric minute (3). Partial trisomy 7 or partial monosomy 7 share some common clinical features, such as mental retardation, growth deficiency and finger abnormalities as well as eye and ear anomalies. In addition, certain patients present variable features, such as triangular face, high frontal hairline, high arched palate, clinodactyly and asymmetry of limbs (4).

Uniparental disomy (UPD), the abnormal inheritance of both chromosomes from only one parent, has been described for different human chromosomes. UPD can be associated with an sSMC as a result of an original trisomy with consecutive trisomic rescue (5).

Congenital adrenal hyperplasia (CAH) is one of the most common autosomal recessive disorders, with an estimated carrier frequency of 1 in 50 (6), and is due to 21-hydroxylase deficiency (21-OHD), one of the enzymes required for the synthesis of cortisol in the adrenal cortex. Impaired 21-hydroxylase activity causes accumulation of steroid precursors, which then flow into biosynthetic pathways unaffected by the enzymatic block, resulting in the production of excess androgens. When androgens are over-secreted from the adrenal glands, the female fetus is virilized (7). Complete enzyme inactivation or low but measurable enzyme activity leads to the salt wasting (SW) form, 3–7% residual enzyme activity leads to the simple virilizing type and more than 30% residual enzyme activity leads to the non-classic type (7). Approximately 75% of patients with classic 21-OHD have the SW form, which is associated with severe impairment of 21-hydroxylation of progesterone and 17-hydroxyprogesterone (17-OHP) (7).

The 21-hydroxylase gene, CYP21 (CYP21A2, OMIM No. 201910), is located on chromosome 6p21.3 within the HLA histocompatibility complex in close proximity to the highly homologous inactive pseudogene, CYP21P (CYP21A1P) (8,9). Recombinations and conversions between CYP21A1P and CYP21A2 result in the generation of dysfunctional CYP21A2 alleles (7,10). Approximately 90% of CYP21A2 mutations are a result of CYP21A1P-derived conversions and recombinations (7,11). CYP21A2 genotyping can be useful in the diagnosis of CAH and can also predict the phenotype in 80–90% of cases (12,13).

In this study, we present a female pseudohermaphroditism/adrenogenital syndrome (AGS) case with a 45% mosaicism for a small supernumerary ring chromosome 7 associated with a homozygous mutation in the CYP21A2 gene.

The karyotype determined by GTG-banding identified was mos 48,XX,+mar1×2[1]/47,XX+mar1[45]/46,XX[54] (Fig. 4). The sSMC was present in 45 of 100 lymphocyte-derived metaphases. FISH excluded the presence of SRY-specific sequences (data not shown). The sSMC was further characterized by molecular cytogenetic studies which revealed a centric minute-shaped chromosome 7 [min(7)(:p11.1->q11.23:)] (Fig. 5). The karyotypes of the mother and the father were 46,XX and 46,XY, respectively.

Sequencing of the CYP21A2 gene revealed the homozygous mutation c.518T>A (p.Ile173Asn) in exon 4 (NM_000500.5; ATG=1). Both parents are heterozygous for that mutation (Fig. 6).

sSMC(7) is very rare and usually small in size. They consist of the centromere and small amounts of euchromatic material, a fact which also applies to our patient. Only 15 patients with sSMC originating from the proximal region of the long arm are described in the literature. Comparing the phenotype of cases reported, delay of speech is often reported (http://www.med.uni-jena.de/fish/sSMC/07.htm#Start07).

The phenotypic spectrum associated with duplication of the 7q11.23 Williams-Beuren Syndrome (WBS) region has previously been delineated. In these patients, the main clinical feature is a moderate to severe expressed language delay (19). By contrast, subjects with deletion of the same interval have good communication competence, with a relative strength in verbal skill. These findings support the idea that this region contains genes that may affect speech performance in a dose-dependent manner. Four genes (CALN1, STX1A, LIMK1 and CYLN), involved in brain development or function, have been identified (20). As was observed in our patient, facial anomalies are non-specific, but some traits are common to both ring chromosome 7 [r(7)] carriers and patients with WBS duplication, and thus they can be associated with 7q proximal region triplication. Our patient shares a prominent forehead, a triangular face, a high nasal bridge, normal eyes, thin lips, short philtrum and normal ears (21) with the other patients. By contrast, other aspects, such as hirsutism, were observed in our case and other cases as well (21). These features should be carefully sought when a new case of r(7) is discovered, since they could represent helpful clues to better delineate the r(7) phenotype.

CAH is found in a wide range of clinical severity ranging from subtle hormone imbalance in adults to severe life-threatening SW in newborns (22). Detection of the underlying mutations in the CYP21A2 gene encoding steroid 21-hydroxylase enzyme is helpful both for confirmation of diagnosis and management of CAH patients (22). Approximately 95% of the mutated alleles in patients with steroid 21-OHD are generated by transfer of DNA sequences from CYP21A1P to CYP21A2 by gene conversion events (23). The remaining 5% of the alleles show new/rare mutations due to random events (24). Most of these mutations are unique to individual families, but some are population-specific (25,26). Different kinds of mutations result in different degrees of enzymatic impairment of P450c21, which result in varied phenotypes of CAH patients. A large number of mutations detected in the CYP21A2 gene have been characterized to prove their clinical relevance and impact on the P450c21 protein. The residual enzyme activity is then measured towards both natural substrates (17-OHP and progesterone) and compared to the wild-type protein. The percentage of the residual enzyme activity is correlated with the clinical phenotype and subsequently mutations are classified as simple virilization, SW or non-classic types of AGS (27–31), with the change c.518T>A being a typical mutation for simple virilization.

To our knowledge, in this study, we present the first report of a female pseudohermaphroditism, classified as simple virilization form of 21-OHD, with an additional min(7(:p11.1->q11.23:) and the homozygous mutation c.518T>A (p.Ile173Asn) in the CYP21A2 gene.