RESUMEN
The genetic diversity of Plasmodium falciparum (P falciparum) infections in humans is implicated in the pathogenesis of malaria. This study provides the first estimate of the genetic diversity and genotype multiplicity of Plasmodium falciparum infection in children with uncomplicated P falciparum malaria in Osogbo, Nigeria. One hundred and one isolates were used for analysis of parasite population polymorphism and genotyped by nested-PCR of merozoite surface protein 2 (MSP2) block 3. Amplicons were obtained for all the 101 genotyped samples in MSP2 PCR with 9 alleles varying in size between 300 and 800 base pair. Thirty-three (31.7%) samples had FC27 allele while 27 (26.7%) had 3D7 allele and 35 (34.7%) had mixed alleles (3D7+FC27). The Multiplicity of Infection (MOI) in the population was 1.6. Children in the age group of > 4-8 years had the highest number of different genotypes in their samples (1.8). The number of MSP2 bands per isolate was lower in the older age group (1.3) but the difference was not statistically significant. Children with parasite density range 5001-10 000 had the highest MOI of 2 while those with parasite density range 1000-5000 had the lowest of 1.5. In conclusion, the present study shows that the field isolates are highly diverse in respect of MSP2 and multiplicity of infection was neither age nor parasite density dependent in the study population.
Asunto(s)
Antígenos de Protozoos/genética , Malaria Falciparum/genética , Plasmodium falciparum/genética , Polimorfismo Genético , Proteínas Protozoarias/genética , Alelos , Animales , Distribución de Chi-Cuadrado , Niño , Preescolar , Femenino , Variación Genética , Genotipo , Humanos , Lactante , Masculino , Nigeria , Reacción en Cadena de la PolimerasaRESUMEN
Deficiency of mannan-binding lectin-associated serine protease 2 (MASP-2) has been associated with infections, whereas high levels appear to increase the risk of inflammatory disorders. Nevertheless, MASP2 haplotypes have been poorly investigated. To overcome haplotyping cost and time consumption, we developed multiplex polymerase chain reactions with sequence-specific primers (PCR-SSP) for 8 single nucleotide polymorphisms (SNPs), reducing the number of necessary reactions from 18 to 7. SNPs were distributed from the promoter to the last exon, and a single PCR-SSP was used for p.D120G. We evaluated the phylogenetic relationships and global distribution of 10 identified haplotypes in 338 Danish individuals with known MASP-2 and MAp19 levels and 309 South Brazilians. Four haplotypes were associated with reduced MASP-2 levels in plasma (lower than 200 ng/mL). Simultaneous association with the highest MASP-2 (over 600 ng/mL) and lowest MAp19 levels (lower than 200 ng/mL) was demonstrated with the intron 9 mutation (Kruskal-Wallis p < 0.0001). Cumulative genotype frequencies predict approximately 0.4% severely deficient and 25% overproducing individuals in both populations. Rapid and low-cost screening of patients with multiplex MASP2 PCR-SSP could be used to identify clinical conditions where MASP-2 (or MAp19) levels may be disease modifying, possibly improving disease outcome through early therapeutic and preventive measures.
Asunto(s)
Enfermedades Autoinmunes/genética , Etnicidad , Infecciones/genética , Serina Proteasas Asociadas a la Proteína de Unión a la Manosa/metabolismo , Empalme Alternativo/genética , Biomarcadores/metabolismo , Brasil/etnología , Dinamarca/epidemiología , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Haplotipos , Ensayos Analíticos de Alto Rendimiento , Humanos , Serina Proteasas Asociadas a la Proteína de Unión a la Manosa/genética , Reacción en Cadena de la Polimerasa Multiplex , Filogenia , Polimorfismo de Nucleótido SimpleRESUMEN
Parasites are accountable for driving diversity within immune gene families. We identified and investigated regulatory single nucleotide polymorphisms (SNPs) in the promoter regions of the tumor necrosis factor receptor superfamily member 18 (TNFRSF18) gene by direct sequencing in a group of male Gabonese individuals exposed to a wide array of parasitic diseases such as malaria, filariasis and schistosomiasis. Two new promoter variants were identified in 40 individuals. Both novel variants were heterozygous and were linked to SNP #rs3753344 (C/T), which has been described. One of the SNP variants (ss2080581728) was close to the general transcription factor site, the TATA box. We further validated these new promoter variants for their allelic gene expression using transient transfection assays. One new promoter variant with two base changes (C/T - ss2080581728/rs3753344) displayed an altered expression of the marker gene. Both novel variants remained less active at the non-induced state in comparison to the major allele. The allele frequencies observed in this study were consistent with data for other African populations. The detection and analysis of these human immune gene polymorphisms contribute to a better understanding of the interaction between host-parasite and expression of Treg activity.
Asunto(s)
Humanos , Masculino , Proteína Relacionada con TNFR Inducida por Glucocorticoide/genética , Interacciones Huésped-Parásitos/genética , Enfermedades Parasitarias/genética , Polimorfismo de Nucleótido Simple/genética , Regiones Promotoras Genéticas/genética , Gabón , Frecuencia de los Genes , Interacciones Huésped-Parásitos/inmunología , Reacción en Cadena de la Polimerasa , Enfermedades Parasitarias/inmunología , TransfecciónRESUMEN
Parasites are accountable for driving diversity within immune gene families. We identified and investigated regulatory single nucleotide polymorphisms (SNPs) in the promoter regions of the tumor necrosis factor receptor superfamily member 18 (TNFRSF18) gene by direct sequencing in a group of male Gabonese individuals exposed to a wide array of parasitic diseases such as malaria, filariasis and schistosomiasis. Two new promoter variants were identified in 40 individuals. Both novel variants were heterozygous and were linked to SNP #rs3753344 (C/T), which has been described. One of the SNP variants (ss2080581728) was close to the general transcription factor site, the TATA box. We further validated these new promoter variants for their allelic gene expression using transient transfection assays. One new promoter variant with two base changes (C/T - ss2080581728/rs3753344) displayed an altered expression of the marker gene. Both novel variants remained less active at the non-induced state in comparison to the major allele. The allele frequencies observed in this study were consistent with data for other African populations. The detection and analysis of these human immune gene polymorphisms contribute to a better understanding of the interaction between host-parasite and expression of Treg activity.
Asunto(s)
Proteína Relacionada con TNFR Inducida por Glucocorticoide/genética , Interacciones Huésped-Parásitos/genética , Enfermedades Parasitarias/genética , Polimorfismo de Nucleótido Simple/genética , Regiones Promotoras Genéticas/genética , Gabón , Frecuencia de los Genes , Interacciones Huésped-Parásitos/inmunología , Humanos , Masculino , Enfermedades Parasitarias/inmunología , Reacción en Cadena de la Polimerasa , TransfecciónRESUMEN
The mannose binding lectin (MBL2) polymorphism is responsible for a common immunodeficiency in the human species. There were suggestions that the MBL2 polymorphism has been under balancing selection, based on the high global frequency of alleles generating MBL deficiency and on the worldwide distribution of diseases negatively associated with them. To describe the distribution of MBL2 allelic haplotypes in Brazilian populations and to discuss the evolution of this polymorphism, we analyzed six South Brazilian populations (152 Guarani Amerindian, 239 Kaingang Amerindian, 107 admixed, Brazilian 32 Afro-Brazilian, 202 Euro-Brazilian and 16 Oriental-Brazilian). Eight haplotypes were observed: MBL2*HYPA, LYQA, LYPA, LXPA, LYPB, LYQC, HYPD, and LYPD. In addition, through sequencing of the promoter and exon 1 from Amerindian and Oriental individuals, three new single-nucleotide polymorphisms (SNPs) were found in the MBL2 promoter region in the Kaingang. Analysis of the sequencing data by neutrality tests (Tajima's D and Fu and Li's D* and F*) revealed no deviation from selective neutrality equilibrium in the Guarani and Kaingang. Significant Fay and Wu's H results are explained by the recent gene flow in these populations. Contrarily to previous thoughts, stochastic evolutionary factors seem therefore to have had a predominant role in shaping the MBL2 polymorphism, at least in the Amerindians.
Asunto(s)
Predisposición Genética a la Enfermedad/epidemiología , Lectina de Unión a Manosa/genética , Polimorfismo Genético , Brasil/epidemiología , Flujo Génico , Haplotipos , Humanos , Desequilibrio de Ligamiento , Reacción en Cadena de la PolimerasaRESUMEN
It was shown that Rhodnius prolixus vitellogenin (Vg) is synthesized as precursors of 205 and 190 kDa. Each Vg subunit is antigenically related to a domain in the precursor molecules. Since Vg has been previously detected in R. prolixus male adults, protein synthesis by fat bodies from 5th instar male nymphs was investigated and no Vg synthesis could be detected. Also, a 6.1 Kb RNA is present in female adults but not in 5th instar male nymphs. Therefore, cDNAs from female adult and 5th instar male fat bodies were used for differential screening of a female fat body cDNA library leading to the isolation of several female specific clones. All the clones hybridizing to the female specific 6.1 Kb RNA species were identical. We also describe the construction of new expression vectors, pGex-A and pGex-B, derived from the previously described plasmid pGex-1N. The new vectors, together with pGex-3X, comprise a set of expression plasmids with cloning sites in all three possible reading frames that give a fusion polypeptide with the glutathione S-transferase. This carrier protein can be cleaved by digestion with factor Xa in all three plasmids; one of the Vg cDNA clones was subcloned in pGex-A. Antibodies affinity purified from the fusion protein Vg/glutathione S-transferase recognized both large Vg subunits, suggesting an antigenic relationship between them. Furthermore, the small Vg subunits were not recognized, indicating that they may be localized at the N-terminal region of Vg precursors.