Disorders of sex development (DSDs) are congenital conditions in which the external appearance of the individual does not coincide with the chromosomal constitution or the gonadal sex. In other words, there is an ambiguous or intermediate condition between the male and female phenotypes of the anatomical sex. These atypical conditions are manifested in several ways, ranging from genital ambiguity to phenotypes that are so attenuated that they can go unnoticed or appear normal. Currently, there is a lack of understanding of the factors responsible for these outcomes; however, they are likely to be conditioned by genetic, hormonal and environmental factors during prenatal and postnatal development. The present study determined the genetic etiology of DSDs in Colombian patients by conventional cytogenetic analysis, FISH and MLPA (for SF1, DAX1, SOX9, SRY and WNT4). A cohort of 43 patients with clinical phenotypes of sex development disorder was used in the present study. Using this multistep experimental approach, a diagnostic percentage of 25.58% was obtained: 17 patients (39.53%) were classified as having gonadal development disorders, the majority of which were ovotesticular disorders with numerical and/or structural alterations of the sex chromosomes, 9 patients (20.93%) were classified as having testicular DSD with a 46,XY karyotype, and 3 patients (6.98%) as having ovarian DSD with a 46,XX karyotype. The remaining 14 patients (32.56%) were classified as ‘other’ since they could not be grouped into a specific class of gonadal development, corresponding to hypospadias and multiple congenital anomalies. These findings highlight the importance of histological and cytogenetic studies in a gonadal biopsy. In 11/43 cases, the multistep experimental protocol presented in the present study yielded etiological or histological findings that could be used to define the medical management of patients with DSDs. In conclusion, for the etiological diagnosis of DSDs, a broad‑spectrum approach that includes endocrinological tests, conventional karyotyping, molecular karyotyping by FISH and, molecular tests is required, in addition to gonadal tissue analyses, to identify genetic alterations.

To analyze the impact of Ascaris lumbricoides infection on the pathogenesis and diagnosis of allergic diseases, new allergens should be identified. We report the identification of a new Ascaris lumbricoides allergen, Asc l 5. The aim of this study was to evaluate the physicochemical and immunological features of the Asc l 5 allergen. We constructed an A. lumbricoides cDNA library and Asc l 5 was identified by immunoscreening. After purification, rAsc l 5 was physicochemically characterized. Evaluation of its allergenic activity included determination of Immunoglobulin E (IgE) binding frequency (in two populations: 254 children and 298 all-age subjects), CD203c based-basophil activation tests (BAT) and a passive cutaneous anaphylaxis (PCA) mouse model. We found by amino acid sequence analysis that Asc l 5 belongs to the SXP/RAL-2 protein family of nematodes. rAsc l 5 is a monomeric protein with an alpha-helical folding. IgE sensitization to rAsc l 5 was around 52% in general population; positive BAT rate was 60%. rAsc l 5 induced specific IgE production in mice and a positive PCA reaction. These results show that Asc l 5 has structural and immunological characteristics to be considered as a new allergen from A. lumbricoides.

Background: Variant curation refers to the application of evidence‐based methods for the interpretation of genetic variants. Significant variability in this process among laboratories affects clinical practice. For admixed Hispanic/Latino populations, underrepresented in genomic databases, the interpretation of genetic variants for cancer risk is challenging.Methods: We retrospectively evaluated 601 sequence variants detected in patients participating in the largest Institutional Hereditary Cancer Program in Colombia. VarSome and PathoMAN were used for automated curation, and ACMG/AMP and Sherloc criteria were applied for manual curation.Results: Regarding the automated curation, 11% of the variants (64/601) were reclassified, 59% (354/601) had no changes in its interpretation, and the other 30% (183/601) presented conflicting interpretations. With respect to manual curation, of the 183 variants with conflicting interpretations, 17% (N = 31) were reclassified, 66% (N = 120) had no changes in their initial interpretation, and 17% (N = 32) remained with conflicting interpretation status. Overall, 91% of the VUS were downgraded and 9% were upgraded.Conclusions: Most VUS were reclassified as benign/likely benign. Since false‐positive and ‐negative results can be obtained with automated tools, manual curation should also be used as a complement. Our results contribute to improving cancer risk assessment and management for a broad range of hereditary cancer syndromes in Hispanic/Latino populations.

Background: Analysis of patients with chromosomal abnormalities, including Turner syndrome and Klinefelter syndrome, has highlighted the importance of X‐linked gene dosage as a contributing factor for disease susceptibility. Escape from X‐inactivation and X‐linked imprinting can result in transcriptional differences between normal men and women as well as in patients with sex chromosome abnormalities.Objective: To identify differentially expressed genes among patients with Turner (45,X) and Klinefelter (46,XXY) syndrome using bioinformatics analysis.Methodology: Two gene expression data sets of Turner (45,X) and Klinefelter syndrome (47,XXY) were obtained from the Gene Omnibus Expression (GEO) database of the National Center for Biotechnology Information (NCBI). Statistical analysis was performed using R Bioconductor libraries. Differentially expressed genes (DEGs) were determined using significance analysis of microarray (SAM). The functional annotation of the DEGs was performed with DAVID v6.8 (The Database for Annotation, Visualization, and Integrated Discovery).Results: There are no genes over‐expressed simultaneously in both diseases. However, when crossing the list of under‐expressed genes for 45,X cells and the list of over‐expressed genes for 47,XXY cells, there are 16 common genes: SLC25A6, AKAP17A, ASMTL, KDM5C, KDM6A, ATRX, CSF2RA, DHRSX, CD99, ZBED1, EIF1AX, MVB12B, SMC1A, P2RY8, DOCK7, DDX3X, eight of which are involved in the regulation of gene expression by epigenetic mechanisms, regulation of splicing processes and protein synthesis.Conclusion: Of the 16 identified as under‐expressed in 45,X cells and over‐expressed in 47,XXY cells, 14 are located in X chromosome and 2 in autosomal chromosome; 8 of these genes are involved in the regulation of gene expression: 5 genes are related to epigenetic mechanisms, 2 in regulation of splicing processes, and 1 in the protein synthesis process. Our results are limited by it being the product of a bioinformatic analysis from mRNA isolated from whole blood, this makes necessary further exploration of the relationships between these genes and Turner syndrome and Klinefelter syndrome in the future.

Introduction: Helminth infections, including those caused by gastrointestinal helminths are characterized by mast cell activation; eosinophilia and high levels of circulating IgE. There is evidence that these antibodies are involved in immunity against parasites. Ascaris lumbricoides has somatic antigens and excretory/secretory products that induce a strong Th2 and specific IgE response; being important examples the protein ABA-1, glutathione transferase (GST) and tropomyosin. Tropomyosinis the major allergenic component of Ascaris extract but other IgE binding components have been detected by one two dimensional SDS-PAGE immunoblotting; some of them homologous to other allergen sources. The identification and characterization of new allergenic molecules from A. lumbricoides is essential for understanding immunity to this helminth and its influence on the evolution of allergic diseases in humans. During the screening of a cDNA library with human IgE antibodies we found a clone coding for cystatin (Al-CPI), a molecule with potential allergenic properties. Objective: To evaluate the ability of recombinant Al-CPI to bind human IgE and induce an allergic response in vitro and in vivo. Methods: We amplify the Ascaris lumbricoides cystatin sequence by PCR strategies. The PCR product was sequenced to confirm correspondence with that reported in GeneBank. A codon-optimized version of the Al-CPI sequence (481 nucleotides) was inserted into the expression vector pQE30 and produced as recombinant protein in E coli; in addition, its inhibitory activity was determined by protease-activity assays using specific colorimetric substrates. The prevalence of specific IgE and IgG4 antibodies against Al-CPI was determined by indirect ELISA, testing 237 sera from a cohort of asthmatic patients naturally exposed to the parasite (range of age: 7 to 70 years old). The ability to activate an allergic response was evaluated ex vivo by basophile activation test (BAT) and histamine release assays (HRA) and in vivo by skin prick test (SPT) in 40 subjects with positive IgE antibody levels to Al-CPI (OD>0.141), ten of them with positive SPT to Ascaris extract. Furthermore, the allergenicity was analyzed in vivo using a passive cutaneous anaphylaxis (PCA) model in BALB/c mice. Results: A recombinant cystatin of 14.2 kDa with potent inhibitory activity of cysteine protease was produced and purified. This protein has 42% identity with the highly allergenic Anisakis simplex cystatin, Ani s 4. The IgE binding frequency to Al-CPI was 64.5% in the group of asthmatic patients sensitized to Ascaris and 56.6% in non-sensitized individuals. When comparing the levels of IgG4, IgE and the ratio IgG4/IgE in Ascaris extract sensitized (n=154) and non-sensitized individuals (n=83), only IgG4 levels were significantly higher in those sensitized (median OD: 0.190 vs 0.177; Mann Whitney test: p= 0.021). In vivo (SPT) or ex vivo (BAT or HRA) functional assays to test the allergenicity of Al-CPI in humans were negative in contrast to the house dust mite allergen Der p 2, which was used as a positive control. In mice, PCA test was also negative. Conclusions: Our findings suggest that Al-CPI induces a strong and frequent IgE response in human exposed to Ascaris lumbricoides but does not induce an allergic reaction, probably, because of the blocking effect of IgG4 antibodies. However, this and other factors underlying the lack of allergenicity of this molecule should be evaluated because it could be a general phenomenon of protective IgE mediated response without adverse allergic effects on the host. The impact of our findings on the relationship of helminthiasis and allergy should be further investigated. Also, it remains to be defined whether this occurs with molecules from other sources.