ABSTRACT
The reproductive mechanism of Giardia intestinalis, considered one of the earliest divergent eukaryotes, has not been fully defined yet. Some evidence supports the hypothesis that Giardia is an exclusively asexual organism with a clonal population structure. However, the high genetic variability, the variation in ploidy during its life cycle, the low heterozygosity and the existence of genes involved in the meiotic-like recombination pathway in the parasite's genome cast doubt on exclusively asexual nature of Giardia. In this work, semiquantitative RT-PCR analysis was used to assess the transcription pattern of three meiosis-like-specific genes involved in homologues recombination: dmc1, hop1 and spo11. The mRNAs were amplified during the parasite's differentiation processes, encystation and excystation, and expression was found at each stage of its life cycle. A semiquantitative assessment also suggests that expression of some of the genes is regulated during encystation process.
Subject(s)
Animals , Genes, Protozoan/genetics , Giardia lamblia/genetics , Meiosis/genetics , Crossing Over, Genetic , Reproduction, Asexual , Reverse Transcriptase Polymerase Chain Reaction , RNA, Messenger , Transcription, GeneticABSTRACT
The reconstruction of Giardia lamblia life cycle in vitro is an excellent tool for the study of the parasite's molecular biology. The present work describes techniques developed that better define parasite differentiation. An encystation protocol is presented along with a method for isolation and purification of the produced cysts. The cyst morphology at the light microscopy level is identical to that of in vivo cysts. A two-dimension protein map obtained by high-resolution electrophoresis indicated that most of the parasite's proteins are acid. Based on this result, the two dimension gel electrophoresis used a pH 4-7 gradient in the first, isoelectric focusing dimension. Differences in protein expression during the stages of encystation were clearly discerned, as well as images of the parasite obtained by light and by transmission electron microscopy that describe the morphological and the ultrastructural changes that occur as the cysts are produced in vitro.