Molecular spectrum of ß-thalassemia mutations in the admixed Venezuelan population, and their linkage to ß-globin gene haplotypes.

In order to establish the spectrum of ß-thalassemia (ß-thal) mutations in the Venezuelan population for the first time, 127 unrelated subjects either with a suspicion of ß-thal trait or with a clinically recognized ß-thal syndrome of different degrees of severity, were studied. DNA from these subjects was analyzed by a polymerase chain reaction (PCR)-based reverse dot-blot method or amplification refractory mutation system (ARMS). Prototype ß-globin gene sequencing of relevant DNA was performed to confirm the mutations. Fifteen different mutations were identified accounting for 92.0% of the mutant alleles explored, revealing a significant genetic heterogeneity at the ß-globin gene locus in this population. The most frequent mutations were codon 39 (C >T) 34.1%, IVS-I-1 (G >A) 11.1%, IVS-I-6 (T > C) 6.6%, IVS-I-110 (G >A) 6.6%, IVS-II-849 (A >G) 6.6%, -88 (C >T) 6.0%, -29 (A >G) 5.2%, followed by the less common IVS-I-5 (G >A) 3.7%, the 1,393 bp deletion 3.0%, IVS-II-1 (G >A) 3.0%, -86 (C >G) 2.2%, IVS-II-1 (G >T) 1.5%, codons 41/42 (-TCTT) 1.5%, IVS-II-745 (C >G) 0.7% and deletional δß-thal 0.7%. Overall, these data demonstrate that the major sources of ß-thal alleles in Venezuela, as expected, are of Mediterranean and African origins. This is the first large study defining the molecular spectrum of ß-thal in the highly admixed population of Venezuela and lays the foundation for genetic counseling as well as implementing comprehensive clinical care programs. Diversity of haplotypes associated with some of the ß-thal mutations can be explained by in situ recombination events in Venezuela.

[Advantages in the use of high performance liquid chromatography technique for screening hemoglobinopathies in Venezuela]. / Ventajas de la técnica de cromatografía líquida de alta presión (HPLC-CE) en el estudio de hemoglobinopatías en Venezuela.

The hemoglobinopathies are a very heterogeneous group of congenital hemolytic anemias, which includes hemoglobin (Hb) variants, thalassemia and hereditary persistence of fetal hemoglobin (HPFH). The aim of this study was to determine the frequency of hemoglobinopathies using the High Performance Liquid Chromatography (HPLC-CE) technique with the beta-thalassemia Short Program of Variant* Bio Rad. Four thousand blood samples from anemic patients from the Laboratorio de Investigación de Hemoglobinas Anormales, Hospital Universitario de Caracas were studied. Twenty six percent of the anemia patients had hemoglobinopathies. The Hb S was the most frequent variant found, followed by the Hb C and Hb D. Also we observed the association of beta thalassemia with Hb S and Hb C. The quantification of the Hb A by HPLC-CE allowed us to classify the double heterozygote Hb S-Beta Thalassemia in Hb S-beta+ Tal Type 1, Hb S-beta+ Tal Type 2, Hb S-beta(0) Thalassemia. The double heterozygote patients with Hb C-Beta thalassemia were also classified. The HPLC-CE is a rapid, reproducible and precise technique. The reliability of HbA2 measurement by HPLC for the detection of beta thalassaemia without any false positive or false negative results is of great advantage. HPLC may be an appropriate method for rapid screening in population surveys for beta thalassemia and hemoglobin variants carriers. Due to the high incidence of cases, in our country this is very important for their clinical management and the genetic and anthropological impact of an early and precise diagnosis.

Ventajas de la técnica de cromatografía líquida de alta presión (HPLC-CE): en el estudio de hemoglobinopatías en Venezuela / Advantages in the use of high performance liquid chromatography technique for screening hemoglobinopathies in Venezuela

Las hemoglobinopatías comprenden un grupo heterogéneo de anemias congénitas, entre las que destacan: las variantes de hemoglobina, las talasemias y la persistencia hereditaria de hemoglobina fetal. Con la técnica de cromatografía líquida de alta presión de intercambio catiónico (HPLC-CE) y utilizando el programa b-thalassemia Short Program® del equipo Variant* Bio Rad, se estudió la frecuencia de hemoglobinopatías en un total de 4000 muestras de pacientes referidos al Laboratorio de Investigación de Hemoglobinas Anormales del Hospital Universitario de Caracas, por presentar problemas de anemia. Se encontró que el 26 por ciento de los pacientes con diagnóstico de anemia, obedece a la presencia de hemoglobinopatías. Entre las detectadas, la Hb S es la variante más frecuente, seguida de las variantes C y D. Además, se observó la presencia de Beta Talasemia y su asociación a las hemoglobinas S y C. Debido a la cuantificación del porcentaje de Hb A por el método de HPLC-CE fue posible clasificar a los pacientes doble heterocigotos Hb S-Beta Talasemia: en Hb S-b+ Tal Tipo 1, Hb S-b+ Tal Tipo 2, Hb S-b0 Talasemia. De la misma manera, fueron clasificados los pacientes doble heterocigotos con Hb C-Beta Talasemia. La técnica de HPLC-CE es sensible y precisa en el estudio de un gran número de muestras, lo cual se logra en muy corto tiempo, tiene un alto poder de resolución y reproducibilidad de los resultados. En todo paciente con anemia hemolítica congénita, la detección de estas patologías es de suma importancia para lograr un adecuado monitoreo, establecer un tratamiento precoz, consejo genético y un manejo terapéutico multidisciplinario