Isoetes louisianensis was discovered in 1972 and listed as an endangered species in 1992. It occurs along shaded streams in southeastern Louisiana and southern Mississippi. Isoetes louisianensis is a fertile tetraploid (2n=44; x=11). To test the hypothesis that I. louisianensis is an allotetraploid, we sequenced the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA of I. louisianensis and aligned it to ITS sequences of the basic diploid species I. engelmannii, I. flaccida, I. melanopoda, and I. valida (all 2n=22), which grow in the vicinity of I. louisianensis. Comparisons of sequence chromatograms indicate that I. engelmannii and I. melanopoda are the most likely progenitors of I. louisianensis. Isoetes engelmannii and I. melanopoda have different nucleotides at ten sites. At seven of these sites I. louisianensis shows additivity for I. engelmannii and I. melanopoda nucleotides. At the other three sites, I. louisianensis has the same nucleotide as either I. engelmannii or I. melanopoda. These molecular results are in agreement with the intermediate megaspore texture and growth habit of I. louisianensis. The boldly cristate-reticulate megaspore texture of I. louisianensis appears to be a blending of the reticulate texture of I. engelmannii megaspores and the obscurely rugulate texture of I. melanopoda megaspores. While I. engelmannii will grow continuously under favorable conditions, I. melanopoda tends to be a spring ephemeral. Isoetes louisianensis appears intermediate with reduced growth in summer and fall. Thus, chromosome number, spore morphology, growth habit, and nucleotide sequences indicate that I. louisianensis is an allotetraploid derivative of I. engelmannii and I. melanopoda.

Key words: allopolyploidy, Isoetes louisianensis, ITS