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The chromosome number and phylogenetic position of the monotypic eusporangiate genus Mankyua (Ophioglossaceae), which shares peculiar morphological characteristics with Ophioglossum s.l. and Helminthostachys, were determined. Meiotic and mitotic chromosome observations of Mankyua show n = 130 and 2n = ca. 260, suggesting that the base number of this genus is x = 130, which is a new finding and the highest base number reported so far for Ophioglossaceae. The molecular phylogenetic trees derived from nucleotide sequence variation of rbcL and matK genes differ in terms of the phylogenetic position of Mankyua. The molecular phylogenetic trees based on combined data sets of rbcL and matK suggest that Mankyua might be the earliest-diverging lineage of Ophioglossaceae.

Several cytotypes (polyploids and aneuploids) have been reported in Lepisorus thunbergianus. The relationships between these cytotypes within the species remain poorly understood. We studied populations in an area where various cytotypes of L. thunbergianus as well as two diploid species, L. angustus and L. onoei, candidate parental species that may be involved in allopolyploid origins of L. thunbergianus polyploids, occur. We determined the ploidy levels of sampled materials by direct chromosome counting and flow cytometry. We elucidated the origins of L. thunbergianus polyploids by analyzing allozyme polymorphisms, and in addition, we examined the occurrence of segmental allopolyploidy by comparing allelic variation between polyploids and their parental diploids. Six cytotypes, i.e. one diploid (2n = 50), two triploid (2n = 75 and 76) and three tetraploid (2n = 100, 101, and 102) cytotypes, were observed in L. thunbergianus, and the two diploid species, L. angustus (2n =52) and L. onoei (2n =50) were confirmed to include a single cytotype each. Allozyme analyses indicated that the tetraploid (2n =100) and hypertetraploid (2n = 102) of L. thunbergianus originated by allopolyploidy between diploid L. thunbergianus (2n = 50) and diploid L. angustus (2n = 52), since the polyploids shared alleles with these two diploids that were unique to each diploid. The allozyme patterns excluded the possibility that L. thunbergianus polyploids originated from L. onoei. The unbalanced heterozygosity and homozygosity found in the tetraploid and the hypertetraploid of L. thunbergianus indicated their segmental allopolyploidy.