The cyprinid fish Alburnus alburnus possesses one of the largest supernumerary chromosomes in all vertebrates. In the present study, amplified fragment length polymorphism analyses (AFLP) and fluorescence in-situ hybridization (FISH) were performed in order to characterize these extraordinary chromosomes in detail. Sequence analysis of the B chromosome-specific DNA revealed a strong homology to a Drosophila Gypsy/Ty3 retrotransposon and also to a medaka (Oryzias latipes) one. The sequence is highly abundant on the B chromosome but undetectable in the normal A chromosome complement. It is also absent from the B chromosome of the closely related species, Rutilus rutilus, suggesting a specific spreading of the mobile element during evolution of the giant supernumerary chromosome within A. alburnus. Meitotic chromosomes were in-situ hybridized with the B chromosome-specific probe, documenting that the additional chromosome behaves as an autopaired ring chromosome in diakineses. Our results suggest that the supernumerary chromosome of A. alburnus is not derived from the normal chromosome complement but has evolved independently.

The Amazon molly, Poecilia formosa, is an all-female fish of hybrid origin which reproduces by gynogenesis, i.e. it depends on sperm of males of closely related species to trigger parthenogenetic development of the embryo. Therefore the offspring is clonal and identical to the mother. In rare cases the exclusion mechanism fails and paternal introgression occurs. This may result either in triploid offspring – if the whole haploid chromosome set of the sperm fuses with the diploid egg nucleus – or in siblings with microchromosomes – if only subgenomic amounts of paternal DNA are included. In one of our diploid, microchromosome-carrying laboratory stocks we observed eight triploid individuals which all developed into males. We investigated the mitotic and meiotic chromosomes, the synaptonemal complex (SC), and sperm production of these males, and compared them to males of the gonochoristic parental species (P. latipinna and P. mexicana) and their hybrids. This comparison revealed that P. formosa males are functional males with reduced effective fertility. They show a deviation from the typical 23 bivalents in the synaptonemal complexes as well as in diakinesis due to the triploid state. They produced offspring but only with gynogenetic Amazon molly females. This shows that the probably aneuploid sperm from P. formosa males can trigger parthenogenetic development of unreduced eggs.

In animals, supernumerary chromosomes and their evolution have mostly been studied in sexual reproducing species. In the present study, for the first time, the natural distribution and stability of supernumerary microchromosomes were investigated in the unisexual fish species Poecilia formosa. Natural habitats throughout the range of P. formosa were screened for the presence of microchromosomes over several years. A high frequency of microchromosomes was found in the Río Purificación river system. Evidence points to the presence of the same microchromosome lineage over many generations. No supernumerary chromosomes were found elsewhere than in the Río Purificación representing a significant difference in the distribution of microchromosome-bearing individuals between the Río Purificación and all other collection sites.

In a microchromosome-carrying laboratory stock of the normally all-female Amazon molly Poecilia formosa triploid individuals were obtained, all of which spontaneously developed into males. A comparison of morphology of the external and internal insemination apparatus and the gonads, sperm ploidy and behaviour, to laboratory-bred F 1 hybrids revealed that the triploid P. formosa males, though producing mostly aneuploid sperm, are partly functional males that differ mainly in sperm maturation and sexual motivation from gonochoristic P. formosa males.

The gynogenetic livebearing Amazon molly (Poecilia formosa) is a sexual parasite that exploits males of closely related species for sperm. This is needed as physiological stimulus for embryo development; however, none of the male's genes are normally incorporated into the genome of the gynogenetic offspring. Mostly diploid individuals were reported from the natural habitats in North-Eastern Mexico and South-Eastern Texas but stable populations of triploids have been reported from the Río Soto la Marina drainage and in the Río Guayalejo in North-Eastern Mexico. Triploidy is the result of defects in the mechanisms that normally clear the host sperm from the ameiotic diploid egg. Triploids also reproduce gynogenetically and their frequencies fluctuate markedly between years, seasons, and localities. To understand the dynamics of this mating system, it is important to understand the relative reproductive success of triploids and diploids. We hypothesize that triploids should have a selective advantage over diploids due to heterosis and/or gene redundancy based on the additional genetic material from the sexual host. However, clonal competition Electronic Supplementary Material The online version of this article (experiments revealed a clear reproductive advantage of diploids competing with triploids. This result contradicts not only our hypothesis but also the stable co-existence of diploids and triploids in natural habitats. Frequency dependent selection, niche partitioning and environmental heterogeneity are discussed as possible explanations.