RESUMO
The factors that characterize Acanthamoeba strains as harmless or potentially pathogenic have not been elucidated. Analysing the in vitro and in vivo parameters of Acanthamoeba samples, including heat tolerance at temperatures close to that of the human body, cytopathic effects, and their ability to cause infections in animals, has been proposed to identify their pathogenic potential. Another promising criterion for differentiating strains is the analysis of their biochemical and immunochemical properties. In this study, a comparative evaluation between clinical and environmental Acanthamoeba isolates was performed on the basis of physiological, morphological, and immunochemical criteria. Crude antigens were used to characterize the protein profiles by electrophoresis and immunize mice to produce polyclonal and monoclonal antibodies. The antibodies were characterized using ELISA, Western blotting, and immunofluorescence techniques. The results obtained with polyclonal antibodies suggest the presence of specific proteins for each studied isolate and co-reactive immunochemical profiles among conserved components. Ten monoclonal antibody clones were obtained; mAb3 recognized 3 out of 4 samples studied. The results of this study may help standardize criteria for identifying and characterizing Acanthamoeba strains. Taken together, our results support the view that a set of features may help differentiate Acanthamoeba species and isolates.
Assuntos
Ceratite por Acanthamoeba/parasitologia , Acanthamoeba/classificação , Poeira/análise , Parasitologia/métodos , Acanthamoeba/imunologia , Acanthamoeba/isolamento & purificação , Acanthamoeba/ultraestrutura , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/imunologia , Anticorpos Antiprotozoários/biossíntese , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/administração & dosagem , Antígenos de Protozoários/imunologia , Antígenos de Protozoários/isolamento & purificação , Western Blotting , Eletroforese , Ensaio de Imunoadsorção Enzimática/métodos , Características da Família , Imunofluorescência , Humanos , Imunização , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Varredura , Proteínas de Protozoários/química , Proteínas de Protozoários/imunologia , Especificidade da EspécieRESUMO
X-linked adrenal hypoplasia congenita (AHC) is caused by mutations in the NR0B1 gene. This gene encodes an orphan member of the nuclear receptor superfamily, DAX1. Ongoing efforts in our laboratory have identified nine novel NR0B1 mutations in X-linked AHC patients (Y81X, 343delG, 457delT, 629delG, L295P, 926-927delTG, 1130delA, 1141-1155del15, and E428X). Two additional families segregate previously identified NR0B1 mutations (501delA and R425T). Sequence analysis of the mitochondrial D-loop indicates that the 501delA family is unrelated through matrilineal descent to our previously analyzed 501delA family.