Role of paraoxonase 1 activity and PON1 gene polymorphisms in sickle cell disease.

Sickle cell disease (SCD) patients often exhibit a dyslipidemic sub-phenotype. Paraoxonase 1 (PON 1) is a serum glycoprotein associated with the high-density lipoproteins cholesterol (HDL-C), and variability in PON1 activity depends on the PON1 genotypes. We investigated the influence of PON1c.192Q > R and PON1c.55L > M polymorphisms on PON1 activity and laboratory parameters and the association between PON1 activity and clinical manifestations in SCD patients. We recruited 350 individuals, including 154 SCD patients and 196 healthy volunteers, which comprised the control group. Laboratory parameters and molecular analyses were investigated from the participants' blood samples. We have found increased PON1 activity in SCD individuals compared to the control group. In addition, carriers of the variant genotype of each polymorphism presented lower PON1 activity. SCD individuals carrying the variant genotype of PON1c.55L > M polymorphism had lower platelet and reticulocyte counts, C-reactive protein, and aspartate aminotransferase levels; in addition to higher creatinine levels. SCD individuals carrying the variant genotype of PON1c.192Q > R polymorphism had lower triglyceride, VLDL-c, and indirect bilirubin levels. Furthermore, we observed an association between PON1 activity history of stroke and splenectomy. The present study confirmed the association between PON1c.192Q > R and PON1c.55L > M polymorphisms and PON1 activity, in addition to demonstrate their effects on markers of dislipidemia, hemolysis and inflammation, in SCD individuals. Moreover, data suggest PON1 activity as a potential biomarker related to stroke and splenectomy.

Cryptosporidium diagnosis in different groups of children and characterization of parasite species.

BACKGROUND: Microscopy and enzyme-linked immunosorbent assay (ELISA) are routinely used for Cryptosporidium diagnosis, without differentiating the parasite species. METHODS: Children's feces were analyzed by modified Ziehl-Neelsen (mZN) and ELISA for Cryptosporidium diagnosis and by polymerase chain reaction-restriction fragment length polymorphism for species identification. RESULTS: Cryptosporidium frequency was 2.6%. The sensitivity and specificity of ELISA were 85.7% and 99.7%, respectively, with excellent concordance with mZN (kappa=0.854). Parasite species were characterized as Cryptosporidium hominis (78.3%), Cryptosporidium felis (17.4%), and Cryptosporidium parvum (4.3%). CONCLUSIONS: Coproantigen ELISA is as efficient as mZN for Cryptosporidium diagnosis. Cryptosporidium genotyping suggests anthroponotic and zoonotic transmission to children.

Cryptosporidium diagnosis in different groups of children and characterization of parasite species

ABSTRACT Background: Microscopy and enzyme-linked immunosorbent assay (ELISA) are routinely used for Cryptosporidium diagnosis, without differentiating the parasite species. Methods: Children's feces were analyzed by modified Ziehl-Neelsen (mZN) and ELISA for Cryptosporidium diagnosis and by polymerase chain reaction-restriction fragment length polymorphism for species identification. Results: Cryptosporidium frequency was 2.6%. The sensitivity and specificity of ELISA were 85.7% and 99.7%, respectively, with excellent concordance with mZN (kappa=0.854). Parasite species were characterized as Cryptosporidium hominis (78.3%), Cryptosporidium felis (17.4%), and Cryptosporidium parvum (4.3%). Conclusions: Coproantigen ELISA is as efficient as mZN for Cryptosporidium diagnosis. Cryptosporidium genotyping suggests anthroponotic and zoonotic transmission to children.

Human infection by Trichostrongylus spp. in residents of urban areas of Salvador city, Bahia, Brazil / Infección humana por Trichostrongylus spp. en residentes de zonas urbanas de la ciudad de Salvador, Bahia, Brasil

Introduction: Nematodes of Trichostrongylus genus are mainly parasites of herbivores, although sporadic human infections have been reported in many countries. Objective: To describe the frequency and seasonality of Trichostrongylus spp. infection in individuals attended at a public clinical laboratory. Materials and methods: Fecal samples of 9,283 individuals were evaluated by spontaneous sedimentation (Lutz) in the Parasitology Laboratory of the Pharmacy College, Federal University of Bahia, Brazil, from January of 2006 to May of 2008. The positive samples for either Trichostrongylus spp. or hookworms were further examined to evaluate the morphometry of nematode eggs. Results: One-hundred and ten patients (1.2%) were confirmed to be infected by Trichostrongylus spp. The positive cases were significantly more frequent in females (1.6%; p <0.05), with higher distribution in the age group between 11-20 years (1.9%), compared to those aged 51-60 (0.8%) and older than 60 years (0.9%)( p <0.05), independent of gender. Trichostrongylus spp. infections were more common from March to May (40 cases) and showed a homogeneous distribution over the other periods of the year (21-25 cases). The hematological analyses of 60 Trichostrongylus -infected patients showed normal levels of eosinophils in most of the positive cases. Conclusions: The data reveal that the occurrence of infection by Trichostrongylus spp. in residents of Salvador is more frequent than those reported in other urban regions and that it is essential to distinguish the parasite from other nematodes in routine parasitological examination.

Introducción. Los nematodos del género Trichostrongylus son parásitos principalmente herbívoros, aunque se hayan descrito en muchos países infecciones humanas esporádicas. Objetivo. Describir la frecuencia y estacionalidad de la infección por Trichostrongylus spp. en individuos atendidos en un laboratorio clínico público. Materiales y métodos. Las muestras fecales de 9.283 individuos fueron analizadas mediante sedimentación espontánea (Lutz) en el Laboratorio de Parasitología de la Facultad de Farmacia de la Universidad Federal de Bahía, Brasil, desde enero de 2006 a mayo de 2008. Las muestras positivas para Trichostrongylus o Ancylostoma fueron reexaminadas para evaluar la morfometría de los huevos de nematodos. Resultados. La infección por Trichostrongylus spp. se confirmó en 110 pacientes (1,2 %), con mayor frecuencia en el sexo femenino (1,6 %; p<0,05). La frecuencia fue mayor en el grupo de edad de 11 a 20 años (1,9 %), independientemente del sexo, en comparación con las personas de 51 a 60 (0,8 %) y con los mayores de 60 años (0,9 %) (p < 0,05). Las infecciones por Trichostrongylus spp. fueron más frecuentes entre marzo y mayo (40 casos) con una distribución homogénea en otros periodos del año (21 a 25 casos). Los análisis hematológicos de 60 pacientes infectados con Trichostrongylus spp. presentaron niveles normales de eosinófilos en la mayoría de los casos positivos. Conclusiones. Los datos revelan que la incidencia de la infección por Trichostrongylus spp. en individuos residentes en Salvador es más frecuente que las descritas en otras áreas urbanas y, por ello, es esencial distinguir dicho parásito de otros nematodos en los exámenes parasitológicos de rutina.

Human infection by Trichostrongylus spp. in residents of urban areas of Salvador city, Bahia, Brazil.

INTRODUCTION: Nematodes of Trichostrongylus genus are mainly parasites of herbivores, although sporadic human infections have been reported in many countries. OBJECTIVE: To describe the frequency and seasonality of Trichostrongylus spp. infection in individuals attended at a public clinical laboratory. MATERIALS AND METHODS: Fecal samples of 9,283 individuals were evaluated by spontaneous sedimentation (Lutz) in the Parasitology Laboratory of the Pharmacy College, Federal University of Bahia, Brazil, from January of 2006 to May of 2008. The positive samples for either Trichostrongylus spp. or hookworms were further examined to evaluate the morphometry of nematode eggs. RESULTS: One-hundred and ten patients (1.2%) were confirmed to be infected by Trichostrongylus spp. The positive cases were significantly more frequent in females (1.6%; p <0.05), with higher distribution in the age group between 11-20 years (1.9%), compared to those aged 51-60 (0.8%) and older than 60 years (0.9%)( p <0.05), independent of gender. Trichostrongylus spp. infections were more common from March to May (40 cases) and showed a homogeneous distribution over the other periods of the year (21-25 cases). The hematological analyses of 60 Trichostrongylus -infected patients showed normal levels of eosinophils in most of the positive cases. CONCLUSIONS: The data reveal that the occurrence of infection by Trichostrongylus spp. in residents of Salvador is more frequent than those reported in other urban regions and that it is essential to distinguish the parasite from other nematodes in routine parasitological examination.

Estudo de polimorfismos em genes de moléculas associadas ao estresse oxidativo na doença falciforme: assciação com dados hematológicos, bioquímicos e fenotípicos / Study polimoorfismos molecules in genes associated with oxidative stress in sickle cell disease: assciação with hematological, biochemical and phenotypic

Indivíduos com doença falciforme (DF) apresentam estresse oxidativo (EO) constante, que é decorrente da deficiência nos sistemas antioxidantes, caracterizados pela produção de espécies reativas do oxigênio (EROS) de origem multifatorial, principalmente geradas pelo processo hemolítico. Objetivo. Investigar polimorfismos relacionados ao estresse oxidativo nos genes HFE da hemocromatose hereditária (c.282C>Y, c.63H>D e c.65S>C); haptoglobina (HP), glutationa S-transferase (GST) (GSTT1 e GSTM1) e paraoxonase (PON1) (c.55L>M e c.192Q>R), associando-os aos dados hematológicos e bioquímicos, aos níveis séricos de vitamina C, paraoxonase 1 e receptor de transferrina e histórico clínico de pacientes com doença falciforme. Casuística e Métodos. A casuística foi composta por 153 indivíduos com DF (HbSS e HbSC) e 196 crianças saudáveis. O perfil de hemoglobinas foi determinado por HPLC. Os polimorfismos nos genes HFE, HP, GST, PON1, a presença da talassemia alfa 3,7Kb e 4,2Kb e dos haplótipos ligados ao grupo de genes da globina beta foram investigados por PCR e PCR-RFLP. A avaliação do perfil lipídico, hepático, função renal, inflamatório e metabolismo do ferro foram realizadas por técnicas espectrofotométricas. Os níveis séricos de vitamina C, receptor de transferrina solúvel e hemeoxigenase 1 total foram detectados por ELISA. Resultados. O estudo de polimorfismos no gene da GST demostrou que o genótipo Mu nulo foi o mais frequente em ambos os grupos estudados; a análise hematológica , nos indivíduos HbSS revelou diferença significante para os parâmetros de leucócitos (p=0,0452), de segmentados neutrófilos (p=0,0224) e o evento de STA (p=0,0423); nos indivíduos com DF foram encontradas diferenças para a uréia (p=0,0288) e ferritina (p=0,0316). No estudo do gene da Haptoglobina nos indivíduos com DF encontrou-se diferença significativa para contagem de reticulócitos (p=0,0167), ferritina sérica (p=0,0493); contagem de linfócitos típicos (p=0.0422) e LDH (p=0,0181); nos HbSS encontrou-se diferença para creatinina (p=0,0178) e ALT (p=0,0127). Para o gene HFE foi encontrada diferença significativa entre os alelos selvagem e mutante para o polimorfismo c.63H>D e contagem de plaquetas (p=0,0311) em indivíduos com DF; para o polimorfismo c.65S>C observou-se diferenças estatisticamente significativas para hemoglobina (p=0,0044) e hematócrito (p=0,0078). Nos HbSC para o polimorfismo c.63H>D observou-se entre os alelos selvagem e mutante diferenças significativas com o VCM (p=0,0032), HCM (p=0,0010) e linfócitos típicos (p=0,0242). Entre os indivíduos HbSS observou-se diferença significativa entre os alelos estudados e plaquetas (p=0,0078). Para a mutação c.65S>C observou-se diferenças significativas para os parâmetros de hemoglobina (p=0,0265) e hematócrito (p=0,0407) nos indivíduos HbSS e hemoglobina (p=0,0051) e hematócrito (p=0,0060) nos indivíduos HbSC. A análise do histórico clínico dos indivíduos com DF e o polimorfismo c.63H>D demonstrou diferença significativa para esplenomegalia, OR=3,38 (0,93 -12,22) e p=0,0402. Nos indivíduos HbSS foi encontrada diferença significativa entre os alelos c.63H>D e infecção, com OR=0,29 (0.07 -1,18) e p=0,0455. Para o gene PON1, a avaliação da atividade da PON1 nos indivíduos DF entre os alelos selvagem e mutante revelou diferença estatisticamente significativa, tendo o alelo selvagem maior atividade da PON1 que alelo mutante; para o polimorfismo PON1c.192Q>R observou-se entre os alelos selvagem e mutante diferenças significativas para as variáveis bioquímicas VLDL-C (p=0,0267) e triglicérides (p=0,0127). Nos HbSC verificou-se diferença estatística significativa para o PON1c.192Q>R e concentração de hemoglobina (p=0,0459), hematócrito (p=0,0225) e contagem de linfócitos típicos (p=0,0364). Para o PON1c.55L>M observou-se diferença significativa com a idade (p=0,0139), plaquetas (p=0,0109), creatinina (p=0,0329) e PCR (p=0,0141). Observou-se associação entre atividade da PON1 e esplenectomia (p=0,001); para hemeoxigenase e ferro sérico (p=0,023); e para vitamina C observamos correlação com colesterol HDL (p=0,037). Há correlação entre glutationa e vitamina C (p=0,035).Conclusões: Os polimorfismos estudados podem estar agindo sinergicamente com a hemoglobina variante S e assim influenciar na gravidade da doença, necessitando de outros estudos para validar estes resultados, uma vez que algumas destas investigações foram realizadas pela primeira vez neste grupo de indivíduos.

Hemoglobinopathies in newborns from Salvador, Bahia, Northeast Brazil.

Hemoglobinopathies are hereditary disorders of the hemoglobin molecule with a high prevalence worldwide. Brazil has a prevalence of 0.1 to 0.3% of newborns with sickle cell anemia and 20.0 to 25.0% of heterozygous alpha2 thalassemia among African Brazilians. In the present study, we investigated the presence of variant hemoglobins and alpha2(3.7 Kb) and alpha2 (4.2 Kb) thalassemia in newborns from Salvador, Bahia, Brazil. Samples of umbilical cord blood from a total of 590 newborns were analyzed, of which 57 (9.8%) were FAS; 36 (6.5%) FAC; one (0.2%) SF; and five (0.9%) FSC. One hundred fourteen (22.2%) newborns had alpha2(3.7 Kb) thalassemia, of whom 101 (19.7%) were heterozygous and 13 (2.5%) homozygous, showing statistical significance for hematological data between newborns with normal alpha genes and alpha2(3.7 Kb) thalassemia carriers. The alpha2(4.2 Kb) thalassemia was not found. Frequencies found in the present study confirm that hemoglobinopathies are a public health problem in Brazil, emphasizing the need for neonatal screening and genetic counseling programs.

Hemoglobinopathies in newborns from Salvador, Bahia, Northeast Brazil

Hemoglobinopatias são alterações hereditárias na molécula de hemoglobina com prevalência mundial elevada. O Brasil apresenta prevalência de 0,1 a 0,3por cento para recém-nascidos com anemia falciforme e freqüência de 20,0 a 25,0 por cento para a ocorrência de heterozigotos da talassemia alfa sobrescrito 2 entre indivíduos afro-descendentes. O presente estudo investigou a presença de hemoglobinas variantes e talassemia alfa 2 subscrito 2 sobrescrito 3.7kb e alfa subscrito 2 sobrescrito 4.2Kb em recém-nascidos de Salvador, Bahia, Brasil. Analisamos o sangue do cordão umbilical de 590 recém-nascidos, sendo 57 (9,8 por cento) com padrão FAS; 36 (6,5 por cento) FAC; um (0,2 por cento) SF e cinco (0,9 por cento) FSC. Cento e catorze (22,2 por cento) apresentaram talassemia alfa 2 subscrito 2 sobrescrito 3.7kb, dos quais 101 (19,7 por cento) foram heterozigotos e 13 (2,5 por cento) homozigotos, mostrando significância estatística para os dados hematológicos entre recém-nascidos com genes a normais e portadores de talassemia alfa 2 subscrito 2 sobrescrito 3.7kb. A talassemia alfa subscrito 2 sobrescrito 4.2Kb não foi encontrada. As freqüências descritas neste trabalho confirmam que as hemoglobinopatias são um problema de Saúde Pública no Brasil, enfatizando a importância dos programas de triagem neonatal e aconselhamento genético.

The beta-globin gene cluster haplotypes in sickle cell anemia patients from Northeast Brazil: a clinical and molecular view.

The beta(S)-globin haplotypes were studied in 78 sickle cell Brazilian patients from Bahia, Northeast Brazil, that has a large population of African origin. Hemoglobin (Hb) profiles were developed by high-performance liquid chromatography (HPLC), and beta(S)-globin gene haplotypes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) techniques. We identified 44 (55.0%) patients with the CAR/Ben (Central African Republic/Benin) genotype, 16 (20.0%) Ben/Ben, 13 (16.2%) CAR/CAR and seven (8.8%) with other genotypes. Analyses of the phenotypes showed clinical differences related only to Hb F levels and blood transfusion therapy; the presence of -alpha(-3.7)-thalassemia (thal) demonstrated statistical significance when associated with hematocrit (p=0.044), MCV (p=0.0007), MCH (p=0.012) and spleen sequestration events. The haplotype diversity found in the present study can be justified by information about the origin of the slave traffic period in Bahia during the 19th century. The specific characteristics described among the Bahian sickle cell patients could be confirmed by increasing the number of patients with specific genotypes and further studies of genetic markers.

C677T polymorphism of the MTHFR gene and variant hemoglobins: a study in newborns from Salvador, Bahia, Brazil.

The C677T polymorphism in the methylenetetrahydrofolate reductase gene (MTHFR) is associated with an increase in total homocysteine serum levels (tHcy), described as a risk factor for cardiovascular disease. Eight hundred forty-three neonates from two different maternity hospitals, one public and another private, in Salvador, Bahia, Brazil were screened for this polymorphism by PCR and RFLP. The T-allele frequency in the total sample was 0.23, and the prevalence rates of heterozygous and homozygous carriers were 36.2% and 5.3%, respectively. The T-allele frequency differed and the T/T genotype was more prevalent at the private maternity hospital. The hemoglobin (Hb) profile was investigated by HPLC in 763 newborns. The frequency of variant Hb was higher at the public than at the private maternity hospital. The association of the C677T polymorphism and the Hb profile was investigated in 683 newborns, showing a relatively high frequency of variant Hbs and the T allele. These data could provide an important basis for further studies focusing on potential risks of vaso-occlusive events in these individuals.

C677T polymorphism of the MTHFR gene and variant hemoglobins: a study in newborns from Salvador, Bahia, Brazil

O polimorfismo C677T no gene da MTHFR tem sido associado ao aumento dos níveis séricos de homocisteína.total (tHcy), descrito como fator de risco para o desenvolvimento de doenças cardiovasculares. Oitocentos e quarenta e três recém-nascidos (RNs), de duas maternidades diferentes, uma pública e a outra privada, em Salvador, Bahia, Brasil foram triados para o polimorfismo C677T por PCR e RFLP. A freqüência do alelo T foi de 0,23 e as prevalências dos genótipos C/T e T/T foram de 36,2 por cento e 5,3 por cento, respectivamente. A freqüência do alelo T diferiu e a prevalência do genótipo T/T foi mais elevada entre os RNs da maternidade privada. O perfil de hemoglobinas (Hb) foi determinado por HPLC em 763 RNs. A freqüência de Hbs variantes foi mais elevada entre os RNs da maternidade pública Tsylla Balbino do que na maternidade privada do Hospital Santo Amaro. A associação do polimorfismo C677T e o perfil de Hbs foram estudados em 683 RNs, apresentando freqüência elevada da coexistência do alelo T e Hb variantes. Estes resultados podem ser utilizados como base para estudos futuros sobre riscos potenciais de eventos vaso-oclusivos nestes indivíduos.