The Mediterranean basin is considered to be one of the 33 mega biodiversity hotspots with the greatest number of endemic plants. This region has been exposed to a prolonged anthropogenic influence rendering its biodiversity prone to a rapid change. The first cytogenetic data are presented for Hedysarum perrauderianum Coss. and Durieu (Fabaceae), an endangered stenoendemic fodder species from Algeria. Karyotype features, heterochromatin organization, physical mapping of rRNA genes and meiotic behaviour were studied in six native populations. The transcriptional rDNA activity was evaluated by nucleoli number. The tetraploid chromosome number 2n = 4x = 32 was found in all populations studied. However, in two roadside populations we observed cells of the same individuals that had 32 and 18 chromosomes. Constitutive heterochromatin is mainly present in intercalary and telomeric positions. Four strong GC-rich DNA bands situated on two pairs of satellite chromosomes are colocalized with 18S-5.8S-26S rDNA loci. Sixteen additional weak CMA bands are also observed in centromeric regions. FISH revealed two loci of both 18S and 5S rDNA, distributed on different chromosome pairs. The similarity of certain chromosome pairs (morphological type of chromosome, constitutive heterochromatin) and presence of univalents, trivalents and tetravalents suggest an autotetraploid origin of this species. One to four heterochromatic B chromosomes were also observed in some populations. The frequencies of different types of chromosome pairing and meiotic abnormalities revealed two groups of populations according to environmental conditions.