Utilizing ethnic-specific differences in minor allele frequency to recategorize reported pathogenic deafness variants.

Ethnic-specific differences in minor allele frequency impact variant categorization for genetic screening of nonsyndromic hearing loss (NSHL) and other genetic disorders. We sought to evaluate all previously reported pathogenic NSHL variants in the context of a large number of controls from ethnically distinct populations sequenced with orthogonal massively parallel sequencing methods. We used HGMD, ClinVar, and dbSNP to generate a comprehensive list of reported pathogenic NSHL variants and re-evaluated these variants in the context of 8,595 individuals from 12 populations and 6 ethnically distinct major human evolutionary phylogenetic groups from three sources (Exome Variant Server, 1000 Genomes project, and a control set of individuals created for this study, the OtoDB). Of the 2,197 reported pathogenic deafness variants, 325 (14.8%) were present in at least one of the 8,595 controls, indicating a minor allele frequency (MAF) > 0.00006. MAFs ranged as high as 0.72, a level incompatible with pathogenicity for a fully penetrant disease like NSHL. Based on these data, we established MAF thresholds of 0.005 for autosomal-recessive variants (excluding specific variants in GJB2) and 0.0005 for autosomal-dominant variants. Using these thresholds, we recategorized 93 (4.2%) of reported pathogenic variants as benign. Our data show that evaluation of reported pathogenic deafness variants using variant MAFs from multiple distinct ethnicities and sequenced by orthogonal methods provides a powerful filter for determining pathogenicity. The proposed MAF thresholds will facilitate clinical interpretation of variants identified in genetic testing for NSHL. All data are publicly available to facilitate interpretation of genetic variants causing deafness.

Clinical and Genetic Characterization of RDH12-Retinal Dystrophy in a South American Cohort.

PURPOSE: To characterize the largest cohort of individuals with retinol dehydrogenase 12 (RDH12)-retinal dystrophy to date, and the first one from South America. DESIGN: Retrospective multicenter international study. SUBJECTS: Seventy-eight patients (66 families) with an inherited retinal dystrophy and biallelic variants in RDH12. METHODS: Review of clinical notes, ophthalmic images, and molecular diagnosis. MAIN OUTCOME MEASURES: Visual function, retinal imaging, and characteristics were evaluated and correlated. RESULTS: Thirty-seven individuals self-identified as Latino (51%) and 34 as White (47%). Sixty-nine individuals (88%) had Leber congenital amaurosis (LCA)/early-onset severe retinal dystrophy. Macular and midperipheral atrophy were seen in all patients from 3 years of age. A novel retinal finding was a hyperautofluorescent ring in 2 young children with LCA. Thirty-nine patients (50%) had subsequent visits, with mean follow-up of 6.8 ± 7.3 (range, 0-29) years. Eight variants (21%) were previously unreported, and the most frequent variant was c.295C>A, p.Leu99Ile, present in 52 alleles of 32 probands. Individuals with LCA homozygous for p.Leu99Ile (31%) had a later age of onset, a slower rate of best-corrected visual acuity decrease, the largest percentage of patients with mild visual impairment, and were predicted to reach legal blindness at an older age than the rest of the cohort. CONCLUSIONS: By describing the largest molecularly confirmed cohort to date, improved understanding of disease progression was possible. Our detailed characterization aims to support research and the development of novel therapies that may have the potential to reduce or prevent vision loss in individuals with RDH12-associated retinal dystrophy. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures.

Phenotypic characterization of retinitis pigmentosa associated with deafness / Caracterización fenotípica de la retinitis pigmentaria asociada a sordera

Introduction: There are several syndromes that associate retinitis pigmentosa with deafness or hearing loss. The most frequent is Usher syndrome, a genetic disorder of autosomal recessive inheritance, which, in some cases, is accompanied by vestibular dysfunction. However, there are cases of families that despite having retinitis pigmentosa associated with deafness, cannot be classified as Usher or other syndromes due to additional findings. Objective: To reassess the phenotypes of 103 families previously diagnosed as possible Usher syndrome and/or retinitis pigmentosa associated with deafness. Materials and methods: We conducted a descriptive and retrospective study by reviewing the medical records of 103 families with a probable clinical diagnosis of Usher syndrome and/or retinitis pigmentosa associated with deafness. Families whose clinical diagnosis did not correspond to the typical Usher syndrome were selected and evaluated ophthalmologically and audiologically. Demographic and clinical variables were analyzed. Results: We selected and then reevaluated 14 families and 55 individuals as they did not correspond to a clinical diagnosis of Usher syndrome; 13.6% of the families initially considered to have typical Usher syndrome were later diagnosed with retinitis pigmentosa associated with deafness, another ocular symptom associated with hearing loss, retinitis pigmentosa, or isolated hearing loss in the same family. Conclusions: Family studies are essential in cases where the symptoms do not match the typical Usher' syndrome. In the cases of retinitis pigmentosa associated with deafness, a correct clinical diagnosis allows for focusing on the molecular analyses to establish a differential diagnosis. The need for nomenclature guidelines on these atypical findings is relevant to aid physicians and researchers in the best approach to these cases.

Introducción. El síndrome de Usher es una alteración genética caracterizada por la asociación de retinitis pigmentaria y sordera. Sin embargo, hay casos con familias en las cuales, a pesar de presentarse dicha asociación, no se puede diagnosticar un síndrome de Usher ni ninguno otro. Objetivo. Reevaluar fenotípicamente a 103 familias con diagnóstico previo de posible síndrome de Usher o retinitis pigmentaria asociada con sordera. Materiales y métodos. Se revisaron las historias clínicas de 103 familias con un posible diagnóstico clínico de síndrome de Usher o retinitis pigmentaria asociada con sordera. Se seleccionaron las familias cuyo diagnóstico clínico no correspondía a un síndrome de Usher típico. Los afectados fueron valorados oftalmológica y audiológicamente. Se analizaron variables demográficas y clínicas. Resultados. Se reevaluaron 14 familias cuyo diagnóstico clínico no correspondía al de síndrome de Usher. De las familias con diagnóstico inicial de síndrome de Usher típico, el 13,6 % recibieron uno posterior de "retinitis pigmentaria asociada con sordera", de "otro síntoma ocular asociado con hipoacusia", o en forma aislada en una misma familia, de "retinitis pigmentaria" o "hipoacusia". Conclusiones. Es fundamental el estudio familiar en los casos en que la clínica no concuerda con el diagnóstico de síndrome de Usher típico. En los pacientes con retinitis pigmentaria asociada con sordera, el diagnóstico clínico acertado permite enfocar los análisis moleculares y, así, establecer un diagnóstico diferencial. Es necesario elaborar guías de nomenclatura en los casos con estos hallazgos atípicos para orientar a médicos e investigadores en cuanto a su correcto manejo.

Diagnostic definition of malattia leventinese in a family from Colombia / Definición diagnóstica en una familia con malattia leventinese en Colombia.

The malattia leventinese is an autosomal dominant inherited disease whose symptoms appear between the second and fourth decades of life. It is characterized by the appearance of drusen located between the retinal pigment epithelium and the Bruch membrane. It is usually associated with low vision and may progress to blindness. The pathogenic variant p.Arg345Trp in the EFEMP1 gene has been associated with this disease. We characterized clinically and molecularly a family with malattia leventinese using a comprehensive approach that involved ophthalmologists, pediatricians, and geneticists. This approach is of great importance since the phenotype of this disease is often confused with acular degeneration. All family members underwent ophthalmological evaluation and DNA extraction from a peripheral blood sample. All exons of the EFEMP1 gene were amplified and sequenced. The pathogenic variant p.Arg345Trp was identified in affected individuals in this family. This is the first report of malattia leventinese in a family with the p.Arg345Trp pathogenic variant in Colombia. The molecular diagnosis of retinal dystrophies is essential to differentiate this type of pathology.

La malattia leventinese es una enfermedad hereditaria autosómica dominante, cuyos síntomas se inician entre la segunda y la cuarta décadas de la vida. Se caracteriza por la aparición de drusas localizadas entre el epitelio pigmentario de la retina y la membrana de Bruch; suele reducir la visión drásticamente y progresar a ceguera. La variante patogénica p.Arg345Trp en el gen EFEMP1 se ha asociado con esta enfermedad. Se presenta aquí la caracterización clínica y molecular de una familia con malattia leventinese mediante un manejo integral que involucró a oftalmólogos, pediatras y genetistas, lo que es de gran importancia, ya que el fenotipo de esta enfermedad suele confundirse con la degeneración macular. A todos los individuos de la familia se les hizo la evaluación oftalmológica con imágenes diagnósticas de retina y extracción de ADN a partir de una muestra de sangre periférica. Todos los exones del gen EFEMP1 se amplificaron y secuenciaron. La variante patogénica p.Arg345Trp se identificó en los individuos afectados. Este es el primer reporte de malattia leventinese en una familia con la variante patogénica p.Arg345Trp en Colombia. El diagnóstico molecular de las distrofias retinianas es fundamental para diferenciar este tipo de enfermedades.

Genetic association with intravitreal ranibizumab response for neovascular age-related macular degeneration in Hispanic population.

BACKGROUND/PURPOSE: Age-related macular degeneration (AMD) is the leading cause of visual impairment in patients over 55 years. Currently, the most common therapies for neovascular AMD (nAMD) are intravitreal antiangiogenics. Studies suggest that genetic factors influence on antiangiogenics therapy outcomes. The purpose of this work was to establish the association between complement factor H (CFH) (Y402H), age-related maculopathy susceptibility 2 (ARMS2) (A69S), and high-temperature requirement factor A1 (HTRA1) (rs11200638) polymorphisms and the response to treatment with ranibizumab in patients with nAMD. METHODS: A cross-sectional study with 61 eyes with nAMD treated with ranibizumab was performed. Association between polymorphisms from CFH, ARMS2, and HTRA1 with the response to treatment was established. RESULTS: The mean age of patients was 76.6 (51-91) years. Only 37.7% of patients had a functional response and 26.2% had an anatomic response. TT polymorphism Y402H from CFH gene was associated with an increased likelihood of functional response to treatment. Otherwise, there was not a statistically significant association between anatomic and functional response to gene polymorphisms rs11200638 from HTRA1 and rs10490924 from ARMS 2. CONCLUSIONS: This study suggests that the response to intravitreal antiangiogenic therapy with ranibizumab was not associated to main polymorphisms from genes HTRA1 and ARMS2. However, it was found that the response to treatment differed according to CFH genotype, suggesting that further investigations are needed to establish if patients with the CC and TC genotype may need to be monitored more closely for disease recurrence than the TT genotype.

Detection of hearing loss in newborns: Definition of a screening strategy in Bogotá, Colombia.

OBJECTIVE: To describe the results from the hearing screening protocol adopted in a Hospital in Colombia emphasizing the importance of performing screening on an outpatient basis, when the newborn is more than 24 h old. METHODS: A prospective study at Hospital Universitario San Ignacio in Bogota, Colombia was carried out, from May 1st, 2016 to Nov 30th, 2017, the study sample included 2.088 newborns examined using transient otoacoustic emissions. RESULTS: We obtained written consent from the parents of 1.523 newborns and 24 individuals (1.6%) failed the first stage of the screening, nine cases unilateral and 15 bilateral. A total of nine neonates (0,6%) failed the second screening test, six cases unilateral and three bilateral. Four (0,3%) did not return to the second test. Our false altered screening rate was 0.7%. CONCLUSIONS: In a developing country with limited human and economic resources, in which newborn early discharge is the norm, a newborn hearing screening program linked to infants' check-ups, that uses otoacoustic emissions after 48 h of life, seems a feasible option compare to the standard US protocol aiming to conduct hearing screening prior to discharge.

A multicenter study on the prevalence and spectrum of mutations in the otoferlin gene (OTOF) in subjects with nonsyndromic hearing impairment and auditory neuropathy.

Autosomal recessive nonsyndromic hearing impairment (NSHI) is a heterogeneous condition, for which 53 genetic loci have been reported, and 29 genes have been identified to date. One of these, OTOF, encodes otoferlin, a membrane-anchored calcium-binding protein that plays a role in the exocytosis of synaptic vesicles at the auditory inner hair cell ribbon synapse. We have investigated the prevalence and spectrum of deafness-causing mutations in the OTOF gene. Cohorts of 708 Spanish, 83 Colombian, and 30 Argentinean unrelated subjects with autosomal recessive NSHI were screened for the common p.Gln829X mutation. In compound heterozygotes, the second mutant allele was identified by DNA sequencing. In total, 23 Spanish, two Colombian and two Argentinean subjects were shown to carry two mutant alleles of OTOF. Of these, one Colombian and 13 Spanish subjects presented with auditory neuropathy. In addition, a cohort of 20 unrelated subjects with a diagnosis of auditory neuropathy, from several countries, was screened for mutations in OTOF by DNA sequencing. A total of 11 of these subjects were shown to carry two mutant alleles of OTOF. In total, 18 pathogenic and four neutral novel alleles of the OTOF gene were identified. Haplotype analysis for markers close to OTOF suggests a common founder for the novel c.2905_2923delinsCTCCGAGCGCA mutation, frequently found in Argentina. Our results confirm that mutation of the OTOF gene correlates with a phenotype of prelingual, profound NSHI, and indicate that OTOF mutations are a major cause of inherited auditory neuropathy.

Screening program for Waardenburg syndrome in Colombia: clinical definition and phenotypic variability.

A screening program to detect Waardenburg syndrome (WS) conducted between 2002 and 2005, among 1,763 deaf individuals throughout Columbia identified 95 affected individuals belonging to 95 families, giving a frequency of 5.38% of WS among the institutionalized deaf population. We confirmed the clinical diagnosis of WS in the 95 propositi and, through the family evaluation, we also identified 45 non-institutionalized affected relatives. Audiologic, ophthalmologic, and genetic studies were performed to confirm the diagnosis. Following the classification of the WS consortium, based on the Waardenburg Index (WI), to define the type of WS. We classified 62.1% of the propositi as WS2 and 37.9% as WS1. We present here the results of the study of clinical manifestations, analyzing the presence, severity, and symmetry of clinical findings among this affected population. Overall, among the 95 propositi, in addition to sensorineural deafness in all, the most frequent features were broad nasal root (58.9%), a first degree relative affected (37.9%), heterochromia irides (36.8%), skin hypopigmentation (31.6%), white forelock (28.0%), intense blue iris (27.4%), synophrys (12.6%), premature graying (10.5%), ptosis of the eyelids (9.5%), and hypoplasia alae nasi (1.1%). The majority of individuals had normal psychomotor development (87%), while the remaining 13% had developmental delay. Among the latter, 9.4% corresponded to WS2 and 3.6% to WS1. Our data confirm an interesting inter- and intrafamilial variability in the phenotypic manifestations as well as extremely variable expression.

Mutational analysis of the LDLR gene in a cohort of Colombian families with familial hypercholesterolemia.

BACKGROUND AND AIMS: Familial hypercholesterolemia (FH) is characterized by elevated serum cholesterol levels due to high low-density lipoprotein (LDL) cholesterol levels. FH is an autosomal dominant genetic disorder and one of the most common dominant hereditary diseases in the world. However, the frequency of mutations in Colombia is unknown. The purpose of this preliminary study was to identify mutations in the LDL receptor (LDLR) gene in a Colombian population with FH. METHODS: The study included 24 families with clinical diagnosis of sure/probable FH. The 18 exons of the LDLR were sequenced by Sanger method. RESULTS: Among 18 variants identified, 3 were known pathogenic mutations and were identified in nine individuals in five unrelated families. Five affected individuals were heterozygous for one mutation each. They were the p.W4X in two, the p.D139G in two and the p.G396D in one. Two affected individuals were homozygous for p.G396D. The variant c.1187-1G > T, which has uncertain significance in FH pathogenesis, was present in all the individuals with the p.D139G mutation. CONCLUSIONS: In total, 18 variants were identified, of which 14 correspond to known nonpathogenic variants. Three pathogenic variants were identified in the LDLR. No pathological mutations were identified in the LDLR in 79% of the study population.

Caracterización fenotípica de la retinitis pigmentaria asociada a sordera / Phenotypic characterization of retinitis pigmentosa associated with deafness

Introducción. El síndrome de Usher es una alteración genética caracterizada por la asociación de retinitis pigmentaria y sordera. Sin embargo, hay casos con familias en las cuales, a pesar de presentarse dicha asociación, no se puede diagnosticar un síndrome de Usher ni ninguno otro. Objetivo. Reevaluar fenotípicamente a 103 familias con diagnóstico previo de posible síndrome de Usher o retinitis pigmentaria asociada con sordera. Materiales y métodos. Se revisaron las historias clínicas de 103 familias con un posible diagnóstico clínico de síndrome de Usher o retinitis pigmentaria asociada con sordera. Se seleccionaron las familias cuyo diagnóstico clínico no correspondía a un síndrome de Usher típico. Los afectados fueron valorados oftalmológica y audiológicamente. Se analizaron variables demográficas y clínicas. Resultados. Se reevaluaron 14 familias cuyo diagnóstico clínico no correspondía al de síndrome de Usher. De las familias con diagnóstico inicial de síndrome de Usher típico, el 13,6 % recibieron uno posterior de "retinitis pigmentaria asociada con sordera" de "otro síntoma ocular asociado con hipoacusia',' o en forma aislada en una misma familia, de "retinitis pigmentaria" o "hipoacusia'.' Conclusiones. Es fundamental el estudio familiar en los casos en que la clínica no concuerda con el diagnóstico de síndrome de Usher típico. En los pacientes con retinitis pigmentaria asociada con sordera, el diagnóstico clínico acertado permite enfocar los análisis moleculares y, así, establecer un diagnóstico diferencial. Es necesario elaborar guías de nomenclatura en los casos con estos hallazgos atípicos para orientar a médicos e investigadores en cuanto a su correcto manejo.

Introduction: There are several syndromes that associate retinitis pigmentosa with deafness or hearing loss. The most frequent is Usher syndrome, a genetic disorder of autosomal recessive inheritance, which, in some cases, is accompanied by vestibular dysfunction. However, there are cases of families that despite having retinitis pigmentosa associated with deafness, cannot be classified as Usher or other syndromes due to additional findings. Objective: To reassess the phenotypes of 103 families previously diagnosed as possible Usher syndrome and/or retinitis pigmentosa associated with deafness. Materials and methods: We conducted a descriptive and retrospective study by reviewing the medical records of 103 families with a probable clinical diagnosis of Usher syndrome and/or retinitis pigmentosa associated with deafness. Families whose clinical diagnosis did not correspond to the typical Usher syndrome were selected and evaluated ophthalmologically and audiologically. Demographic and clinical variables were analyzed. Results: We selected and then reevaluated 14 families and 55 individuals as they did not correspond to a clinical diagnosis of Usher syndrome; 13.6% of the families initially considered to have typical Usher syndrome were later diagnosed with retinitis pigmentosa associated with deafness, another ocular symptom associated with hearing loss, retinitis pigmentosa, or isolated hearing loss in the same family. Conclusions: Family studies are essential in cases where the symptoms do not match the typical Usher' syndrome. In the cases of retinitis pigmentosa associated with deafness, a correct clinical diagnosis allows for focusing on the molecular analyses to establish a differential diagnosis. The need for nomenclature guidelines on these atypical findings is relevant to aid physicians and researchers in the best approach to these cases.

[Phenotypic and molecular characterization of a Colombian family with phenylketonuria].

INTRODUCTION: Phenylketonuria is a metabolic disorder characterized by severe neurological involvement and behavioral disorder, whose early diagnosis enables an effective treatment to avoid disease sequelae, thus changing the prognosis. Objective: To characterize a family with phenylketonuria in Colombia at clinical, biochemical and molecular levels. Materials and methods: The population consisted of seven individuals of a consanguineous family with four children with suggestive symptoms of phenylketonuria. After signing an informed consent, blood and urine samples were taken for colorimetric tests and high performance liquid and thin layer chromatographies. DNA extraction and sequencing of the 13 exons of the PAH gene were performed in all subjects. We designed primers for each exon with the Primer 3 software using automatic sequencing equipment Abiprism 3100 Avant. Sequences were analyzed using the SeqScape, v2.0, software. Results: We described the clinical and molecular characteristics of a Colombian family with phenylketonuria and confirmed the presence of the mutation c.398_401delATCA. We established a genotype-phenotype correlation, highlighting the interesting clinical variability found among the affected patients despite having the same mutation in all of them. Conclusions: Early recognition of this disease is very important to prevent its neurological and psychological sequelae, given that patients reach old age without diagnosis or proper management.

Definición diagnóstica en una familia con malattia leventinese en Colombia / Diagnostic definition of malattia leventinese in a family from Colombia

Resumen La malattia leventinese es una enfermedad hereditaria autosómica dominante, cuyos síntomas se inician entre la segunda y la cuarta décadas de la vida. Se caracteriza por la aparición de drusas localizadas entre el epitelio pigmentario de la retina y la membrana de Bruch; suele reducir la visión drásticamente y progresar a ceguera. La variante patogénica p.Arg345Trp en el gen EFEMP1 se ha asociado con esta enfermedad. Se presenta aquí la caracterización clínica y molecular de una familia con malattia leventinese mediante un manejo integral que involucró a oftalmólogos, pediatras y genetistas, lo que es de gran importancia, ya que el fenotipo de esta enfermedad suele confundirse con la degeneración macular. A todos los individuos de la familia se les hizo la evaluación oftalmológica con imágenes diagnósticas de retina y extracción de ADN a partir de una muestra de sangre periférica. Todos los exones del gen EFEMP1 se amplificaron y secuenciaron. La variante patogénica p.Arg345Trp se identificó en los individuos afectados. Este es el primer reporte de malattia leventinese en una familia con la variante patogénica p.Arg345Trp en Colombia. El diagnóstico molecular de las distrofias retinianas es fundamental para diferenciar este tipo de enfermedades.

Abstract The malattia leventinese is an autosomal dominant inherited disease whose symptoms appear between the second and fourth decades of life. It is characterized by the appearance of drusen located between the retinal pigment epithelium and the Bruch membrane. It is usually associated with low vision and may progress to blindness. The pathogenic variant p.Arg345Trp in the EFEMP1 gene has been associated with this disease. We characterized clinically and molecularly a family with malattia leventinese using a comprehensive approach that involved ophthalmologists, pediatricians, and geneticists. This approach is of great importance since the phenotype of this disease is often confused with macular degeneration. All family members underwent ophthalmological evaluation and DNA extraction from a peripheral blood sample. All exons of the EFEMP1 gene were amplified and sequenced. The pathogenic variant p.Arg345Trp was identified in affected individuals in this family. This is the first report of malattia leventinese in a family with the p.Arg345Trp pathogenic variant in Colombia. The molecular diagnosis of retinal dystrophies is essential to differentiate this type of pathology.

Transient evoked oto-acoustic emission screening in newborns in Bogotá, Colombia: a retrospective study.

OBJECTIVE: The aim of this study was to investigate the characteristics and performance of transient evoked oto-acoustic emission (TEOAE) hearing screening in newborns in Colombia, and analyze all possible variables and factors affecting the results. MATERIALS AND METHODS: An observational, descriptive and retrospective study with bivariate analysis was performed. The study population consisted of 56,822 newborns evaluated at the private institution, PREGEN. TEOAE testing was carried out as a pediatric hearing screening test from December 2003 to March 2012. The database from PREGEN was revised, and the protocol for evaluation included the same screening test performed twice. Demographic characteristics were recorded and the newborn's background was evaluated. Basic statistics of the qualitative and quantitative variables, and statistical analysis were obtained using the chi-square test. RESULTS: Of the 56,822 records examined, 0.28% were classed as abnormal, which corresponded to a prevalence of 1 in 350. In the screened newborns, 0.08% had a major abnormality or other clinical condition diagnosed, and 0.29% reported a family history of hearing loss. A prevalence of 6.7 in 10,000 was obtained for microtia, which is similar to the 6.4 in 10,000 previously reported in Colombia (database of the Latin-American Collaborative Study of Congenital Malformations - ECLAMC). Statistical analysis demonstrated an association between presenting with a major anomaly and a higher frequency of abnormal results on both TEOAE tests. CONCLUSIONS: Newborns in Colombia do not currently undergo screening for the early detection of hearing impairment. The results from this study suggest TEOAE screening tests, when performed twice, are able to detect hearing abnormalities in newborns. This highlights the need to improve the long-term evaluation and monitoring of patients in Colombia through diagnostic tests, and to provide tests that are both sensitive and specific. Furthermore, the use of TEOAE screening is justified by the favorable cost: benefit ratio demonstrated in many countries worldwide.

The importance of fundus eye testing in rubella-induced deafness.

OBJECTIVE: The purpose of this study was to establish a new approach to improve detection of deafness due to rubella. METHODS: Colombian institutes for the deaf were visited by a medical team to perform in all enrolled individuals an ophthalmological examination with emphasis in fundus eye by a retina specialist. In cases where ocular alterations compatible with CRS were found, a medical interview by a clinical geneticist analyzing pre-and postnatal history and a thorough medical examination was done. RESULTS: A total of 1383 deaf institutionalized individuals were evaluated in 9 Colombian cities in the period of 2005 to 2006, finding a total of 463 positive cases for salt-and-pepper retinopathy (33.5%), in which rubella could be the etiology of deafness. Medellin, Cartagena, Bucaramanga and Barranquilla were the cities with the highest percentage of Congenital rubella, corresponding to 22.8% of analyzed population. The analysis performed on cases in which reliable prenatal history was obtained in a second appointment (n=88) showed association between positive viral symptoms during pregnancy and salt-and-pepper retinopathy in 62.5% of cases, while both (retinopathy and viral symptoms) were absent in 29.5% of cases; showing a correlation in 92% of cases. CONCLUSIONS: The frequency of deafness by rubella obtained by this study is significantly high compared with previous Colombian studies and with international reports. It was possible to correlate the antecedent of symptoms during pregnancy with the presence of salt-and-pepper retinopathy in this deaf population when reliable prenatal history was available, therefore eye testing with emphasis in fundus examination is a good indicator of rubella induced deafness. We propose a new approach in the search of deafness causes, based on a thorough ophthalmologic examination in all deaf people.

[Mutational frequencies in usherin(USH2A gene) in 26 Colombian individuals with Usher syndrome type II]. / Frecuencia de mutaciones en el gen de la usherina (USH2A) en 26 individuos colombianos con síndrome de Usher, tipo II.

INTRODUCTION: Usher syndrome is a disorder characterized by progressive retinitis pigmentosa, prelingual sensory hearing loss and vestibular dysfunction. It is the most frequent cause of deaf-blindness in humans. Three clinical types and twelve genetic subtypes have been characterized. Type II is the most common, and among these cases, nearly 80% have mutations in the USH2A gene. OBJECTIVE: The aim of the study was to establish the mutational frequencies for the short isoform of USH2A gene in Usher syndrome type II. MATERIALS AND METHODS: Twenty-six Colombian individuals with Usher syndrome type II were included. SSCP analysis for 20 exons of the short isoform was performed and abnormal patterns were sequenced. Sequencing of exon 13 of the USH2A gene was performed for all the individuals because the most frequent mutation is located in this exon. RESULTS: The most frequent mutation was c.2299delG, identified in the 27% (n=8) of the sample. The second mutation, p.R334W, showed a frequency of 15%. A new variant identified in the 5'UTR region, g.129G>T, was present in 1 individual (4%). Four polymorphisms were identified; one of them is a new deletion in exon 20, first reported in this study. CONCLUSIONS: Mutations in the usherin short isoform were identified in 38% of a sample of 26 USH2 cases. Molecular diagnosis was established in 7 of the 26.

Allele-specific impairment of GJB2 expression by GJB6 deletion del(GJB6-D13S1854).

Mutations in the GJB2 gene, which encodes connexin 26, are a frequent cause of congenital non-syndromic sensorineural hearing loss. Two large deletions, del(GJB6-D13S1830) and del(GJB6-D13S1854), which truncate GJB6 (connexin 30), cause hearing loss in individuals homozygous, or compound heterozygous for these deletions or one such deletion and a mutation in GJB2. Recently, we have demonstrated that the del(GJB6-D13S1830) deletion contributes to hearing loss due to an allele-specific lack of GJB2 mRNA expression and not as a result of digenic inheritance, as was postulated earlier. In the current study we investigated the smaller del(GJB6-D13S1854) deletion, which disrupts the expression of GJB2 at the transcriptional level in a manner similar to the more common del(GJB6-D13S1830) deletion. Interestingly, in the presence of this deletion, GJB2 expression remains minimally but reproducibly present. The relative allele-specific expression of GJB2 was assessed by reverse-transcriptase PCR and restriction digestions in three probands who were compound heterozygous for a GJB2 mutation and del(GJB6-D13S1854). Each individual carried a different sequence variant in GJB2. All three individuals expressed the mutated GJB2 allele in trans with del(GJB6-D13S1854), but expression of the GJB2 allele in cis with the deletion was almost absent. Our study clearly corroborates the hypothesis that the del(GJB6-D13S1854), similar to the larger and more common del(GJB6-D13S1830), removes (a) putative cis-regulatory element(s) upstream of GJB6 and narrows down the region of location.

Caracterización fenotípica y molecular de una familia colombiana con fenilcetonuria / Phenotypic and molecular characterization of a Colombian family with phenylketonuria

Introducción. La fenilcetonuria es un trastorno metabólico caracterizado por un compromiso neurológico grave y por alteraciones del comportamiento. Su diagnóstico temprano permite establecer un tratamiento efectivo que evita las secuelas y modifica el pronóstico. Objetivo. Caracterizar a una familia con fenilcetonuria en Colombia, a nivel clínico, bioquímico y molecular. Materiales y métodos. Se estudió una población de siete individuos de una familia consanguínea en la que cuatro hijos presentaban signos clínicos sugestivos de fenilcetonuria. Una vez firmado el consentimiento informado, se tomaron muestras de sangre y orina para las pruebas colorimétricas, la cromatografía de capa fina y la cromatografía líquida de alta eficacia. Se extrajo el ADN y se secuenciaron los 13 exones del gen PAH de todos los sujetos estudiados. Se diseñaron iniciadores para cada exón con el programa Primer 3; la secuenciación automática se hizo con el equipo Abiprism 3100 Avant y, el análisis de las secuencias, con el programa SeqScape v2.0. Resultados. Se describieron las características clínicas y moleculares de una familia colombiana con fenilcetonuria en la que se identificó la mutación c.398_401delATCA; se presentó una correlación fenotipo-genotipo con una interesante variabilidad clínica entre los afectados, a pesar de tener la misma mutación. Conclusiones. Es importante el reconocimiento temprano de esta enfermedad para evitar sus secuelas neurológicas y psicológicas, pues los pacientes llegan a edades avanzadas sin diagnóstico ni tratamiento adecuados.

Introduction: Phenylketonuria is a metabolic disorder characterized by severe neurological involvement and behavioral disorder, whose early diagnosis enables an effective treatment to avoid disease sequelae, thus changing the prognosis. Objective: To characterize a family with phenylketonuria in Colombia at clinical, biochemical and molecular levels. Materials and methods: The population consisted of seven individuals of a consanguineous family with four children with suggestive symptoms of phenylketonuria. After signing an informed consent, blood and urine samples were taken for colorimetric tests and high performance liquid and thin layer chromatographies. DNA extraction and sequencing of the 13 exons of the PAH gene were performed in all subjects. We designed primers for each exon with the Primer 3 software using automatic sequencing equipment Abiprism 3100 Avant. Sequences were analyzed using the SeqScape, v2.0, software. Results: We described the clinical and molecular characteristics of a Colombian family with phenylketonuria and confirmed the presence of the mutation c.398_401delATCA. We established a genotype-phenotype correlation, highlighting the interesting clinical variability found among the affected patients despite having the same mutation in all of them. Conclusions: Early recognition of this disease is very important to prevent its neurological and psychological sequelae, given that patients reach old age without diagnosis or proper management.

Frecuencia de mutaciones en el gen de la usherina (USH2A) en 26 individuos colombianos con síndrome de Usher, tipo II / Mutational frequencies in usherin (USH2A gene) in 26 Colombian individuals with Usher syndrome type II

Introducción. El síndrome de Usher se caracteriza por hipoacusia neurosensorial congénita, retinitis pigmentaria y disfunción vestibular. Es la causa más frecuente de sordo-ceguera en el mundo. Se divide en tres tipos clínicos y doce subtipos genéticos. El tipo II es la forma más común y cerca de 80 % de los casos corresponden al subtipo 2 del síndrome de Usher. Objetivo. Establecer la frecuencia de mutaciones en la isoforma corta del gen USH2A en individuos colombianos con síndrome de Usher, tipo II. Materiales y métodos. Se estudiaron 26 individuos colombianos con diagnóstico clínico de síndrome de Usher, tipo II. Se hizo análisis de SSCP para los 20 exones que codifican para la isoforma corta y se secuenciaron los patrones anormales. Además, se secuenció el exón 13 en todos los individuos, ya que allí se encuentra la mutación más frecuente de este gen. Resultados. La mutación más frecuente es la c.2299delG, correspondiente al 27 % de la población. La segunda mutación identificada es la p.R334W, con una frecuencia de 15 %. Se identificó un nuevo cambio, el g.129G>T,en la región 5’UTR del gen, correspondiente al 4 % de la población. Se identificaron cuatro cambios polimórficos, uno de ellos es una deleción nueva identificada en el exón 20. Conclusiones. Se logró establecer que, al menos, 38 % de la población analizada con síndrome de Usher, tipo II, presenta alguna mutación en la isoforma corta del gen de la usherina. El diagnóstico molecular se logró establecer en el 23 %.

Introduction. Usher syndrome is a disorder characterized by progressive retinitis pigmentosa, prelingual sensory hearing loss and vestibular dysfunction. It is the most frequent cause of deaf-blindness in humans. Three clinical types and twelve genetic subtypes have been characterized. Type II is the most common, and among these cases, nearly 80% have mutations in the USH2A gene. Objective. The aim of the study was to establish the mutational frequencies for the short isoform of USH2A gene in Usher syndrome type II. Materials and methods. Twenty-six Colombian individuals with Usher syndrome type II were included. SSCP analysis for 20 exons of the short isoform was performed and abnormal patterns were sequenced. Sequencing of exon 13 of the USH2A gene was performed for all the individuals because the most frequent mutation is located in this exon. Results. The most frequent mutation was c.2299delG, identified in the 27% (n=8) of the sample. The second mutation, p.R334W, showed a frequency of 15%. A new variant identified in the 5’UTR region, g.129G>T, was present in 1 individual (4%). Four polymorphisms were identified; one of them is a new deletion in exon 20, first reported in this study. Conclusions. Mutations in the usherin short isoform were identified in 38% of a sample of 26 USH2 cases. Molecular diagnosis was established in 7 of the 26.

Mitochondrial DNA studies show asymmetrical Amerindian admixture in Afro-Colombian and Mestizo populations.

The origin of the African populations that arrived on the Colombian coasts at the time of the Spanish conquest and their subsequent settlement throughout the country and interaction with Amerindian and Spanish populations are features that can be analyzed through the study of mitochondrial DNA (mtDNA) markers. For this purpose, the present study investigates the admixture between these populations by analyzing the markers defining the main (A, B, C, D) and minor (X) founder haplogroups in Native Americans, the principal African haplogroup (L), and additional generic markers present in Caucasian (I, J, K, H, T, U, V, W) and minor African lineages (L3). As part of an interdisciplinary research program (the Expedición Humana, furthered by the Universidad Javeriana and directed by J.E. Bernal V.), 159 Afro-Colombians from five populations in which they are the majority and 91 urban Mestizos were studied. No Amerindian haplogroups (A-D, X) were detected in 81% of the Afro-Colombians. In those samples with Amerindian lineages (average 18.8%, with a range from 10% to 43%), haplogroup B predominated. When analyzed for the presence of African haplotypes, Afro-Colombians showed an overall frequency of 35.8% for haplogroup L mtDNAs, although with broad differences between populations. A few Afro-Colombian samples (1.9%) had mutations that have not been described before, and might therefore be considered as previously unsampled African variants or as new mutations arising in the American continent. Conversely, in Mestizos less than 22% of their mtDNAs belonged to non-Amerindian lineages, of which most were likely to be West Eurasian in origin. Haplogroup L mtDNAs were found in only one Mestizo (1.1%), indicating that, if present, admixture with African women would bring in other, rarer African lineages. On the other hand, in an accompanying paper (Keyeux et al. 2002) we have shown that Amerindians from Colombia have experienced little or no matrilineal admixture with Caucasians or Africans. Taken together, these results are evidence of different patterns of past ethnic admixture among Africans, Amerindians, and Spaniards in the geographic region now encompassing Colombia, which is also reflected in much of the region's cultural diversity.

Possible migration routes into South America deduced from mitochondrial DNA studies in Colombian Amerindian populations.

Mitochondrial DNA haplotype studies have been useful in unraveling the origins of Native Americans. Such studies are based on restriction site and intergenic deletion/insertion polymorphisms, which define four main haplotype groups common to Asian and American populations. Several studies have characterized these lineages in North, Central, and South American Amerindian, as well as Na Dene and Aleutian populations. Siberian, Central Asian, and Southeast Asian populations have also been analyzed, in the hope of fully depicting the route(s) of migration between Asia and America. Colombia, a key route of migration between North and South America, has until now not been studied. To resolve the current lack of information about Colombian Amerindian populations, we have investigated the presence of the founder haplogroups in 25 different ethnic groups from all over the country. The present research is part of an interdisciplinary program, Expedición Humana, fostered by the Universidad Javeriana and Dr. J. E. Bernal V. The results show the presence of the four founder A-D Amerindian lineages, with varied distributions in the different populations, as well as the presence of other haplotypes in frequencies ranging from 3% to 26%. These include some unique or private polymorphisms, and also indicate the probable presence of other Asian and a few non-Amerindian lineages. A spatial structure is apparent for haplogroups A and D, and to a lesser extent for haplogroup C. While haplogroup A and D frequencies in Colombian populations from the northwestern side of the Andes resemble those seen in Central American Amerindians more than those seen in South American populations, their frequencies on the southeastern side more closely resemble the bulk of South American frequencies so far reported, raising the question as to whether they reflect more than one migration route into South America. High frequencies of the B lineage are also characteristic of some populations. Our observations may be explained by historical events during the pre-Columbian dispersion of the first settlers and, later, by disruptions caused by the European colonization.