## 4 Discussion  
In this study of 267 patients with DSD, of the 93 patients that completed all applicable tests, a diagnosis was obtained for 87 (94%) of the patients. A total of 174 patients did not pursue genetic testing and remained etiology unknown following karyotyping, _SRY_ testing, and _CYP21A2_ strip testing. Karyotype testing established the diagnosis in 15% of patients, most of whom presented abnormalities of the sex chromosomes. A value of basal 17-hydroxyprogesterone above the threshold of 2 ng/mL or of stimulated 17-hydroxyprogesterone above 10 ng/mL indicated a deficiency of 21-hydroxylase, and genetic testing of _CYP21A2_ confirmed the diagnosis in 17 out of 39 patients (44% of _CYP21A2_ tested patients). SNP array analysis was particularly useful in DSD patients who presented associated signs (syndromic DSD). Of patients tested by SNP array, 20% received a final etiologic diagnosis, and almost all of them, with one exception, presented syndromic DSD (thispercentage was similar between 46,XX and 46,XY DSD). Gene panel sequencing, which included DSD-associated genes (known and candidate) established or strongly suggested the etiologic diagnosis in 11 of 22 patients tested (50%). The androgen receptor gene mutations were most observed in the study group, but variants in some genes less commonly associated with DSD were also observed, such as those in the _GATA4_ gene. Regarding the percentage of chromosomal abnormalities observed in patients with DSD, a similar result (around 15%) has been found by other studies, and most of these abnormalities involve the sex chromosomes [18; 19]. Thus, karyotype has a role not only in establishing the first step in a pathogenetic algorithm (in 46,XX or 46,XY DSD) but also in observing an etiology in a number of cases (in sex chromosome DSD). The karyotype also provides advantages for identifying mosaicisms, translocations, and X chromosome structural variants, which in some situations are not easily suggested using chromosomal microarray [13]. Concerning the CNVs observed in the study group, the percentage of pathogenic CNVs and uniparental disomy was 20%, similar to that found in other studies, which ranged from 15-20% [18; 20; 21; 22; 23]. One study found that chromosomal microarray is similar to classical cytogenetics when identifying [PERSON] syndrome [24]. However, chromosomal microarray analysis has advantages over karyotype regarding the identification of cryptic Y chromosome material in patients with [PERSON] syndrome [13]. For example, 1q21 deletion (involving _RBM8A_ gene), identified in a 46,XX patient, is a described etiology in Mayer-Rokitansky-Kuster-Hauser syndrome, which is present in some patients with this BP2-BP3 deletion [25]. Another patient was diagnosed with 21-hydroxylase deficiency due to homozygous deletion identified by SNP array testing. The presence of maternal uniparental disomy of 15 and uniparental disomy of chromosome 7 in three of the 35 patients studied by chromosomal microarray provided useful information, as the phenotype of these disorders (Prader-Willi or Russell-Silver syndromes) includes signs of DSD usually associated with other findings that are less obvious. Concerning the other pathogenic CNVs or VUS described in the study, a clear association was not found between genes included in these regions and DSD; these CNVs or genes have been more frequently associated with neurologic development than that of other systems or organs. High-throughput sequencing was found to be the most effective in cases of DSD patients due to its high percentages of positive results. The findings of high-throughput sequencing indicated etiologic and pathogenetic mechanisms, which could be very important for designing optimal therapy and achieving a better prognosis. Of the study's patients, 50% had a pathogenic or likely pathogenic variant responsible for their clinical development, and a similar percentage was observed in a large international cohort of DSD patients [26].