Intraspecific Variation and Phylogenetic Relationships in the Genus Entamoeba as Revealed by Riboprinting

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Abstract
Eighty-seven isolates of amebae assigned to the genus Entamoeba have been studied by riboprinting (restriction enzyme polymorphism analysis of polymerase chain reaction amplified small subunit ribosomal RNA genes). Twenty-four distinct patterns were obtained, most of which corresponded to previously described species. In three species (Entamoeba coli, Entamoeba gingivalis and Entamoeba moshkovskii) intraspecific variation was detected that led to the grouping of isolates into ‘ribodemes’ (populations of amebae that share the same riboprint pattern). The riboprint data were used to estimate genetic distances among and within species for the construction of phylogenetic trees based on parsimony and distance analyses. The trees obtained with the two methods are largely congruent. In some cases the estimated distances between species were greater than the upper limit recommended for the fragment comigration method of analysis indicating unusually deep branches within this genus. However, it appears that those species producing cysts with eight nuclei, those producing cysts with one nucleus, and those producing cysts with four nuclei form morphologically based groups that are supported by the riboprint data. The oral parasite Entamoeba gingivalis, which does not encyst, clusters with the third group indicating secondary loss of this ability.