Abstract
The amoeboflagellate, Naegleria gruberi strain NEG, is a fascinating organism that undergoes a process of cell differentiation when starved, taking it from an amorphous amoeba to a swimming flagellate. NEG has the potential to be a useful model for answering important biological questions, but it lacks genetic tools. Although strain NEG, studied thoroughly by the Fulton lab for over half a century, has not been observed to have a sexual cycle, the genome provides strong evidence that it does. Another species, Naegleria minor strain WT043, shows behavior that suggests sexuality, as does the Naegleria species PNG2, here defined as N. indonesiensis. During differentiation, strain PNG2 undergoes 2-3 cell divisions over the span of 8 hours, during which it forms small flagellates. These small flagellates are not culturable, but when incubated with bacteria some become culturable. We believe that this process of differentiation involves meiosis and gametogenesis. In this scenario, the small flagellates are “gametes” and the cells that regain culturability are “zygotes.” In order to establish a genetic system in PNG2, we planned to use an isozyme electrophoresis assay in which we “mated” PNG2 by inducing differentiation, and then we examined the isozyme band pattern of putative recombinants (amoebae that have regained culturability) compared with that of “parental” PNG2 on a polyacrylamide protein gel, after staining for specific enzymes. These zymograms were examined for inheritance of banding patterns. The inherited banding patterns represent a genetic inheritance for the expression of a specific isozyme and thus should provide evidence of genetic exchange in PNG2. We are using this method in order to demonstrate mating and explore genetic variability in the species of Naegleria that show this apparent sexual cycle.