Variation in chromosome numbers, CMA bands and 45S rDNA sites in species of Selaginella (Pteridophyta)

Background and Aims Selaginella is the largest genus of heterosporous pteridophytes, but karyologically the genus is known only by the occurrence of a dysploid series of n = 7-12, and a low frequency of polyploids. Aiming to contribute to a better understanding of the structural chromosomal variability of this genus, different staining methods were applied in species with different chromosome numbers. Methods The chromosome complements of seven species of Selaginella were analysed and, in four of them, the distribution of 45S rDNA sites was determined by fluorescent in situ hybridization. Additionally, CMA/DA/DAPI and silver nitrate staining were performed to investigate the correlation between the 45S rDNA sites, the heterochromatic bands and the number of active rDNA sites. Key Results The chromosome numbers observed were 2n = 18, 20 and 24. The species with 2n = 20 exhibited chromosome complement sizes smaller and less variable than those with 2n = 18. The only species with 2n = 24, S. convoluta, had relatively large and asymmetrical chromosomes. The interphase nuclei in all species were of the chromocentric type. CMA/DA/DAPI staining showed only a weak chromosomal differentiation of heterochromatic bands. In S. willdenowii and S. convoluta eight and six CMA(+) bands were observed, respectively, but no DAPI(+) bands. The CMA(+) bands corresponded in number, size and location to the rDNA sites. In general, the number of rDNA sites correlated with the maximum number of nucleoli per nucleus. Ten rDNA sites were found in S. plana (2n = 20), eight in S. willdenowii (2n = 18), six in S. convoluta (2n = 24) and two in S. producta (2n = 20). Conclusions The remarkable variation in chromosome size and number and rDNA sites shows that dramatic karyological changes have occurred during the evolution of the genus at the diploid level. These data further suggest that the two putative basic numbers of the genus, x = 9 and x = 10, may have arisen two or more times independently.