Maintenance of variability in tardigrades: parthenogens vs amphimictics in Richtersius coronifer

Parthenogenetic and amphimictic populations within a same morphospecies are frequently reported in non-marine tardigrades. A same morphospecies can have diploid bisexual population(s) reproducing by amphimixis, and polyploid (rarely diploid) unisexual population(s) carrying out apomixis. We assess the breeding system of two Italian populations and one Swedish population of the eutardigrade Richtersius coronifer evaluating sex-ratios, analyzing karyotypes and examining genetic structure using allozyme markers. The populations differed in sex ratio and reproductive modes. One Italian population was bisexual and amphimictic, while the other two were unisexual and parthenogenetic. Data from several years show no evidence of cyclical parthenogenesis. All populations were diploid with the chromosome number (2n = 12) and had bivalents in the oocytes. The chromosome pairing found in the parthenogen oocytes is evidence of a meiotic maturation. The presence of chiasmata in oocytes suggests that recombination exists, even though we do not know how the diploid chromosome number is restored. With regard to the genetic structure, we analyzed 14 loci both in the amphimictic and in the parthenogenetic populations. Only three loci (GPI, IDH and PGM) were polymorphic. The amphimictic population had a degree of genetic variability (mean heterozygosity higher than 0.25) higher than that of the parthenogenetic population. In fact, in all female populations, allele frequencies at all 3 loci deviated from Hardy-Weinberg expectations due to lack of heterozygosity, in accordance with the cytological results evidencing an automictic parthenogenesis. GPI locus was represented by 6 alleles in the amphimictic populations and only by one in the automictic populations, but IDH and PGM were represented by 4 alleles both in the amphimictic and in the automictic populations, evidencing a maintained (or acquired) variability. We cannot exclude that the recent derivation of the asexual populations from few lineages (demographic bottlenecks) could explain the loss of genetic variation, in terms of number of alleles per locus, even though in populations affected by recent demographic bottleneck we expect to observe a relative heterozygosity excess due to the fact that allelic diversity decreases more rapidly than heterozygosity. We found high genetic differentiation between populations of R. coronifer. The observed value of Fst (0.281, i.e. 28% of the total genetic variation is due to differences between populations) is above the threshold (0.25) usually considered indicative of high differentiation among populations. Genetic divergence is easy to explain in this species: differences in reproductive mode, fragmented habitat, patchy distribution and the ensuing difficulty in crossing from one moss fragment to another (i.e. low gene flow and high likelihood of population bottlenecks) are probably all acting on R. coronifer populations.