Resumo Objetivo: Descrever os resultados obtidos em um programa de triagem neonatal após sua implementação e avaliar os perfis clínicos e moleculares de casos confirmados e suspeitos de hiperplasia adrenal congênita. Métodos: Foi feito um estudo transversal. Recém-nascidos com suspeita da doença devido aos altos níveis de 17-alfa-hidroxiprogesterona e ajustados pelo peso ao nascer foram selecionados. A hiperplasia adrenal congênita clássica (forma perdedora de sal e forma virilizante simples) foi diagnosticada por um aumento nos níveis de 17-alfa-hidroxiprogesterona confirmado no reteste, avaliação clínica e genótipo determinado com o uso do ensaio SNaPshot e amplificação multiplex de sondas dependente de ligação. Resultados: Após 24 meses, 15 casos clássicos de hiperplasia adrenal congênita foram diagnosticados em 217.965 recém-nascidos, com uma incidência estimada de 1:14.531. De 132 pacientes, sete não clássicos e 14 heterozigotos foram submetidos à triagem para mutações no gene CYP21A2 e 96 pacientes apresentaram resultados falso-positivos com CYP21A2 do tipo selvagem. No reteste, níveis aumentados de 17-alfa-hidroxiprogesterona foram encontrados em pacientes com hiperplasia adrenal congênita clássica e mostraram correlação significativa com HAC clássica relacionada ao genótipo. As mutações mais frequentes foram IVS2-13A/C>G, seguidas de deleção gênica ou eventos de rearranjo na forma clássica. Em casos de doenças não clássicas e heterozigose, a mutação p.Val282Leu foi a mais comum. Conclusões: Os resultados ressaltam a eficácia da triagem neonatal para a hiperplasia adrenal congênita no sistema público de saúde e indicam que a estratégia adotada foi adequada. A segunda coleta de amostras, juntamente com a genotipagem dos casos suspeitos, ajudou a diagnosticar adequadamente os casos graves e mais leves e diferenciá-los de pacientes com resultado falso-positivo.

Abstract Objective: To describe the results obtained in a neonatal screening program after its implementation and to assess the clinical and molecular profiles of confirmed and suspicious congenital adrenal hyperplasia cases. Methods: A cross-sectional study was conducted. Newborns with suspected disease due to high 17-hydroxyprogesterone levels and adjusted for birth weight were selected. Classical congenital adrenal hyperplasia (salt-wasting and simple virilizing forms) was diagnosed by an increase in 17-hydroxyprogesterone levels as confirmed in the retest, clinical evaluation, and genotype determined by SNaPshot and multiplex ligation-dependent probe amplification. Results: After 24 months, 15 classic congenital adrenal hyperplasia cases were diagnosed in a total of 217,965 newborns, with an estimated incidence of 1:14,531. From 132 patients, seven non-classical and 14 heterozygous patients were screened for CYP21A2 mutations, and 96 patients presented false positives with wild type CYP21A2. On retest, increased 17-hydroxyprogesterone levels were found in classical congenital adrenal hyperplasia patients and showed significant correlation with genotype-related classical genital adrenal hyperplasia. The most frequent mutations were IVS2-13A/C>G followed by gene deletion or rearrangement events in the classical form. In non-classical and heterozygous diseases, p.Val282Leu was the most common mutation. Conclusions: The results underscore the effectiveness of congenital adrenal hyperplasia neonatal screening in the public health system and indicate that the adopted strategy was appropriate. The second sample collection along with genotyping of suspected cases helped to properly diagnose both severe and milder cases and delineate them from false positive patients.

Abstract Sickle cell hemoglobin is the result of a mutation at the sixth amino acid position of the beta (β) globin chain. The HBB*S gene is in linkage disequilibrium with five main haplotypes in the β-globin-like gene cluster named according to their ethnic and geographic origins: Bantu (CAR), Benin (BEN), Senegal (SEN), Cameroon (CAM) and Arabian-Indian (ARAB). These haplotypes demonstrated that the sickle cell mutation arose independently at least five times in human history. The distribution of βS haplotypes among Brazilian populations showed a predominance of the CAR haplotype. American populations were clustered in two groups defined by CAR or BEN haplotype frequencies. This scenario is compatible with historical records about the slave trade in the Americas. When all world populations where the sickle cell gene occurs were analyzed, three clusters were disclosed based on CAR, BEN or ARAB haplotype predominance. These patterns may change in the next decades due to recent migrations waves. Since these haplotypes show different clinical characteristics, these recent migrations events raise the necessity to develop optimized public health programs for sickle cell disease screening and management.

Alpha thalassemia has not been systematically investigated in Brazil. In this study, 493 unrelated individuals from the southernmost Brazilian state of Rio Grande do Sul were screened for deletional forms of α-thalassemia. One hundred and one individuals had microcytic anemia (MCV < 80 fL) and a normal hemoglobin pattern (Hb A2 < 3.5% and Hb F < 1%). The subjects were screened for -α3.7,-α4.2,-α20.5, -SEA and -MED deletions but only the -α3.7 allele was detected. The -α3.7 allele frequency in Brazilians of European and African ancestry was 0.02 and 0.12, respectively, whereas in individuals with microcytosis the frequency was 0.20. The prevalence of α-thalassemia was significantly higher in individuals with microcytosis than in healthy individuals (p = 0.001), regardless of their ethnic origin. There were also significant differences in the hematological parameters of individuals with -α3.7/αα, -α3.7/α3.7 and β-thalassemia trait compared to healthy subjects. These data suggest that α-thalassemia is an important cause of microcytosis and mild anemia in Brazilians.

Glucose-6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) deficiency is one of the most common human enzymopathies throughout the world. Although most affected individuals are asymptomatic, there is a risk of neonatal jaundice and acute hemolytic anemia which can be triggered by infection, some pharmaceuticals and, in older individuals, eating fava beans. We characterized the molecular basis of G6PDH deficiency in a sample of 348 adults from Porto Alegre (population about 1.5 million), the capital of the southernmost Brazilian state of Rio Grande do Sul. Genomic DNA was extracted from peripheral blood leukocytes. We studied the three G6PDH mutations that appear to be the most frequent in Southern Brazil, the G202A and A376G A minus (A-) variants and the C563T Mediterranean (Med) variant. From July 2004 to October 2005, 348 patients (162 Females plus 186 males, age range 0 to 82 years) from Porto Alegre were referred to our laboratory for G6PDH analysis, 36 (9.7%) of which showed deficient G6PDH activity. These 36 patients and 34 randomly-selected non-deficient control individuals were submitted to molecular analysis which revealed a predominance of G6PDH A- allele among the deficient patients. The prevalence of the G6PDH A- variant agrees with its distribution among the ethnic groups that colonized RS, especially those of African, Portuguese, Spanish, and Italian origin.

As leucemias agudas caracterizam-se pela proliferação clonal e pelo bloqueio maturativo das células hematopoéticas, com substituição difusa da medula óssea por células neoplásicas. A leucemia mielóide aguda (LMA) é um grupo heterogêneo de doenças clonais do tecido hematopoético, que acomete predominantemente idosos acima de 60 anos de idade. A LMA apresenta oito subtipos distintos morfologicamente: LMA M0 a M7. Os métodos diagnósticos para identificação da LMA e classificação dos subtipos são baseados em critérios morfológicos, citoquímicos e de imunofenotipagem, acrescidos de análise genética. Além de ser importante para a diferenciação do tipo da linhagem da leucemia, se mielóide (LMA) ou linfóide (LLA), o diagnóstico é também de grande importância para identificar a leucemia bifenotípica aguda (BAL). O objetivo deste trabalho foi realizar uma revisão bibliográfica sobre LMA, dando ênfase aos métodos laboratoriais utilizados para a sua identificação e diferenciação.

The acute leukemias are characterized by the clonal proliferation and maturative blockage of hematopoietic cells, with diffuse substitution of the bone marrow by neoplasic cells. The acute myeloid leukemia (AML) is a heterogeneous group of clonal disease in the hematopoietic tissue and predominantly affects people older than 60. The AML has eight morphologically different subtypes: AML M0 to M7. The diagnostic methods for identification of AML and subtypes classification are based on morphological, cytochemical and immunophenotyping patterns, besides genetic and molecular analyses. The diagnosis of leukemia is important to the lineage differentiation in AML or ALL and also for the identification of biphenotypic acute leukemia (BAL). The aim of this study was to perform a bibliographic review of AML, giving emphasis on laboratory methods useful for its identification and differentiation.