RESUMEN
BACKGROUND: Lutzomyialongipalpis (Diptera: Psychodidae) is the major vector of Leishmania (Leishmania) infantum and thus plays a crucial role in the epidemiology of American visceral leishmaniasis (AVL). This vector is the best studied species of sand fly in the Neotropical region. Many studies claim that this vector is in fact a species complex; however there is still no consensus regarding the number of species that belong into this complex or the geographical distribution of sibling species. The aim of the present study was to analyze the genetic relationships within Lu. longipalpis populations in the state of Mato Grosso do Sul (MS), Brazil. METHODOLOGY/PRINCIPAL FINDINGS: We collected 30 Lu. longipalpis (15 females and 15 males) from five localities (Campo Grande, Três Lagoas, Aquidauana, Miranda and Bonito) and 30 Lu. Cruzi from Corumbá, totaling 180 sandflies from MS, and 30 Lu. longipalpis from Estrela de Alagoas, state of Alagoas (AL), Northeast Brazil. We show that eight previously described microsatellite loci were sufficient in distinguishing Lu. longipalpis from Lu. Cruzi, which is a closely related species, and in differentiating between Lu. longipalpis collected in MS versus Estrela de Alagoas. Analyses of the genotypes revealed introgression between sympatric Lu. longipalpis and Lu. Cruzi. CONCLUSIONS/SIGNIFICANCE: Our findings support the hypothesis of cryptic species within the Lu. longipalpis complex. Furthermore, our data revealed introgression between Lu. longipalpis and Lu. cruzi. This phenomenon should be further investigated to determine the level and incidence of hybridization between these two species. We also demonstrated that microsatellite markers are a powerful tool for differentiating sand fly populations and species. The present study has elucidated the population structure of Lu. longipalpis in MS and, by extension, the Neotropical Lu. longipalpis complex itself.
Asunto(s)
Repeticiones de Microsatélite/genética , Psychodidae/genética , Animales , Femenino , Masculino , Filogenia , Psychodidae/clasificaciónRESUMEN
We investigated the occurrence of Leishmania infantum chagasi in Didelphis albiventris opossums at a wild animal rehabilitation center in the city of Campo Grande, Brazil. A total of 54 opossums were tested for L. i. chagasi infection in peripheral blood and bone marrow samples. The samples were analyzed by direct examination, culturing in a specific medium, and polymerase chain reaction-restriction fragment length polymorphism. Leishmania i. chagasi DNA was detected by polymerase chain reaction-restriction fragment length polymorphism in 11 (20.37%) animals. A total of 81.81% of positive opossums were captured in areas of known visceral leishmaniasis transmission. These results suggest a role for D. albiventris in the urban transmission of visceral leishmaniasis.
Asunto(s)
ADN Protozoario/aislamiento & purificación , Didelphis/parasitología , Leishmania infantum/patogenicidad , Leishmaniasis Visceral/epidemiología , Leishmaniasis Visceral/veterinaria , Animales , Brasil/epidemiología , ADN Protozoario/genética , Femenino , Leishmania infantum/genética , Masculino , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , Población UrbanaRESUMEN
The nuclear lamina is a structure that lines the inner nuclear membrane. In metazoans, lamins are the primary structural components of the nuclear lamina and are involved in several processes. Eukaryotes that lack lamins have distinct proteins with homologous functions. Some years ago, a coiled-coil protein in Trypanosoma brucei, NUP-1, was identified as the major filamentous component of its nuclear lamina. However, its precise role has not been determined. We characterized a homologous protein in Trypanosoma cruzi, TcNUP-1, and identified its in vivo DNA binding sites using a chromatin immunoprecipitation assay. We demonstrate for the first time that TcNUP-1 associates with chromosomal regions containing large non-tandem arrays of genes encoding surface proteins. We therefore suggest that TcNUP-1 is a structural protein that plays an essential role in nuclear organization by anchoring T. cruzi chromosomes to the nuclear envelope.
Asunto(s)
ADN Protozoario/metabolismo , Proteínas de Unión al ADN/análisis , Proteínas Nucleares/genética , Proteínas Protozoarias/genética , Trypanosoma cruzi/química , Inmunoprecipitación de Cromatina , Clonación Molecular , ADN Protozoario/química , Proteínas de Unión al ADN/metabolismo , Membrana Nuclear/química , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Proteínas Protozoarias/química , Proteínas Protozoarias/metabolismo , Trypanosoma cruzi/genética , Trypanosoma cruzi/metabolismoRESUMEN
A molecular epidemiological study was performed with Babesia bigemina isolates from five geographical regions of Brazil. The genetic analysis was done with random amplification of polymorphic DNA (RAPD), repetitive extragenic palindromic elements-polymerase chain reaction (REP-PCR) and enterobacterial repetitive intergenic consensus sequences-polymerase chain reaction (ERIC-PCR) that showed genetic polymorphism between these isolates and generated fingerprinting. In RAPD, ILO872 and ILO876 primers were able to detect at least one fingerprinting for each B. bigemina isolate. The amplification of B. bigemina DNA fragments by REP-PCR and ERIC-PCR gave evidence for the presence in this haemoprotozoan of the sequences described previously in microorganisms of the bacterial kingdom. For the first time it was demonstrated that both techniques can be used for genetic analysis of a protozoan parasite, although the ERIC-PCR was more discriminatory than REP-PCR. The dendogram with similarity coefficient among isolates showed two clusters and one subcluster. The Northeastern and Mid-Western isolates showed the greatest genetic diversity, while the Southeastern and Southern isolates were the closest. The antigenic analysis was done through indirect fluorescent antibody technique and Western blotting using a panel of monoclonal antibodies directed against epitopes on the merozoite membrane surface, rhoptries and membrane of infected erythrocytes. As expected, the merozoite variable surface antigens, major surface antigen (MSA)-1 and MSA-2 showed antigenic diversity. However, B cell epitopes on rhoptries and infected erythrocytes were conserved among all isolates studied. In this study it was possible to identify variable and conserved antigens, which had already been described as potential immunogens. Considering that an attenuated Babesia clone used as immunogen selected populations capable of evading the immunity induced by this vaccine, it is necessary to evaluate more deeply the cross-protection conferred by genetically more distant Brazilian B. bigemina isolates and make an evaluation of the polymorphism degree of variable antigens such as MSA-1 and MSA-2
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Animales , Femenino , Bovinos , Genética , Polimorfismo GenéticoRESUMEN
Plasmid pMV158 encodes a 45 amino acid transcriptional repressor, CopG, which is involved in copy number control. A new procedure for overproduction and purification of the protein has been developed. The CopG protein thus obtained retained its ability to specifically bind to DNA and to repress its own promoter. Purified CopG protein has been crystallized using the sitting-drop vapor diffusion method. The crystals, belonging to orthorhombic space group C222(1) (cell constants a = 67.2 A, b = 102.5 A, c = 40.2 A), were obtained from a solution containing methylpentanediol, benzamidine and sodium chloride, buffered to pH 6.7. Complete diffraction data up to 1.6 A resolution have been collected. Considerations about the Matthews parameter account for the most likely presence of three molecules in the asymmetric unit (2.27 A3/Da).
Asunto(s)
ADN Helicasas , Proteínas de Unión al ADN , Plásmidos , Proteínas/genética , Proteínas Represoras/genética , Transactivadores , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Cristalografía por Rayos X , ADN Bacteriano , Escherichia coli/genética , Vectores Genéticos , Datos de Secuencia Molecular , Estructura Secundaria de Proteína , Proteínas/química , Proteínas/aislamiento & purificación , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Represoras/química , Proteínas Represoras/aislamiento & purificación , Difracción de Rayos XRESUMEN
Streptococcus pneumoniae genetic systems designed for isolation of plasmid mutants with copy-up phenotypes have been developed. The target plasmids have the pLS1 replicon, and two different strategies have been followed: (i) selection of clones exhibiting augmented resistance to antibiotics, or (ii) obligatory co-existence of incompatible plasmids. We have isolated 23 plasmid mutants exhibiting increased number of copies. All the mutations corresponded to four different alleles of the copG gene of plasmid pLS1. These strategies could be used with other plasmids.
Asunto(s)
Dosificación de Gen , Plásmidos/genética , Streptococcus pneumoniae/genética , Secuencia de Aminoácidos , Secuencia de Bases , Genes Bacterianos/genética , Datos de Secuencia Molecular , Mutación/genéticaRESUMEN
Competent cells of Bacillus subtilis are able to bind but not to take up DNA in magnesium-free medium. Wall-membrane complexes, but not cell walls, bind homologous DNA. Binding of DNA to membrane vesicles suggests that DNA-receptor sites are located at the membrane level. On the basis of competition experiments with homologous and heterologous DNAs, in cells and in membrane vesicles, different types of DNA receptors are detected.