Maintenance of pure hybridogenetic water frog populations: Genotypic variability in progeny of diploid and triploid parents

Abstract

An intriguing outcome of hybridisation is the emergence of clonally and hemiclonally reproducing hybrids, that can sustain, reproduce, and lead to the emergence of polyploid forms. However, the maintenance of diploid and polyploid hybrid complexes in natural populations remains unresolved. We selected water frogs from the Pelophylax esculentus complex to study how diploid and triploid hybrids, which reproduce hemiclonally via hybridogenesis, are maintained in natural populations. During gametogenesis in diploid hybrids, one of the parental genomes is eliminated, and the remaining genome is endoreplicated. In triploid hybrids, the single-copy genome is typically eliminated, while genome endoreplication does not occur. To investigate how diploid and triploid hybrid frogs reproduce in populations without parental species, we crossed these hybrid animals from two separate pure hybrid populations located in Poland. Using cytogenetic analysis of tadpoles that emerged from the crosses, we established which gametes were produced by parental hybrids. The majority of hybrid females and hybrid males produced one type of gamete with the P. ridibundus genome. However, in both studied populations, approximately half of the diploid and triploid hybrids simultaneously produced gametes with different genome compositions and ploidy levels, specifically, the P. ridibundus and P. lessonae genomes, as well as diploid gametes with genomes of both parental species. Triploid hybrid males and females mostly produced haploid gametes with the P. lessonae genome; however, gametes with the P. ridibundus genome have also been observed. These results suggest that not all hybrids follow the classical hybridogenetic reproduction program and reveal a significant level of alterations in the gametogenesis pathways. In addition, we found a variable survival rate of particular progeny genotypes when we crossed hybrid females with different males suggesting the important role of postzygotic barriers on the maintenance of pure hybrid systems. We suggest that the observed variability in produced gametes and the different survival rate of the progeny with certain genotypes is crucial for the existence of pure hybrid systems.

Genome elimination from the germline cells in diploid and triploid male water frogs Pelophylax esculentus

“…This repetitive sequence was observed in the pericentromeric region of 5 large and one heterobranchial medium-small chromosome (no. 8) (Ragghianti et al, 1995(Ragghianti et al, , 2004, while others reported its presence in all 13 chromosomes of P. ridibundnus, but with different intensity of signals (7 chromosomes exhibit strong signals, 6 chromosomes exhibit weak signal) (Marracci et al, 2011;Dedukh et al, 2020b;Dedukh et al, 2022b). Moreover, we detected signals in diploid and triploid P. esculentus in interphase nuclei of somatic and germ cells which significantly improved our analysis.…”

“…Hybridogenetic gametogenesis usually occurs in females, except of diploid fish Hypseleotris spp (Schmidt et al, 2011;Majtánová et al, 2021) and triploid green toad Bufotes pseudoraddei baturae (Stöck et al, 2002(Stöck et al, , 2012 where both sexes eliminate one of the genomes. The same process evidently also occurs in male P. esculentus, although it was proven in most cases indirectly by analysis of genome composition in progeny resulting from experimental crosses between parental species and hybrids, as evidenced by classic studies of Berger (Berger, 1968(Berger, , 1973(Berger, , 1976 and others (Dedukh et al, 2017;Dedukh et al, 20220b;Dedukh et al, 2022b). The other evidence comes from studies on genome compositions of spermatozoa detected by flow cytometry that allowed distinguishing genomes on the basis of species-specific difference in amount of DNA in P. lessonae and P. ridibundus (Mazin and Borkin, 1979;Vinogradov et al, 1990;Biriuk et al, 2016).…”

“…Whole genomes of both parental species were labeled either directly or indirectly using Nick Translation Kit (Roche Diagnostics) according to the protocol of Jenkins and Hasterok (Jenkins and Except for whole-genome probes, we also used FISH with centromeric satellite RrS1 probe, which enables clear distinction between P. ridibundus and P. lessonae chromosomes (Ragghianti et al, 1995;Ragghianti et al, 2004;Marracci et al, 2011). Probe was labeled with biotin-11-dUTP or digoxigenin-11-dUTP by PCR amplification in a C1000 Thermal Cycler (Bio-Rad, United States) according to Dedukh et al (Dedukh et al, 2019;Dedukh et al, 2022b).…”

“…Triploid hybrids emerge in rare cases when hybrid individuals produce diploid gametes (RL, LL or RR) (Uzzell, Berger and Günther, 1975;Berger, Roguski and Uzzell, 1978;Christiansen, 2009;Pruvost, Hoffmann and Reyer, 2013;Dedukh et al, 2017;Dedukh et al, 2022a). On the other hand, triploids produce haploid gametes and transmit to progeny the genome which is double-copied (e.g., RRL produces R gametes) (Christiansen, 2009;Dedukh et al, 2015;Dedukh et al, 2022b). A commonly accepted model of hybridogenesis in water frogs assumes that the genome that remains reduplicates before meiotic recombination and forms identical (clonal) copies that eventually are transmitted to clonal gametes.…”

“…We argue that only G (but not SSC) are competent in chromosome rejection. The elimination takes place during interphase via micronuclei budding off from gonocyte nuclei (Ogielska, 1994;Chmielewska et al, 2018) and, probably, in the course of mitosis via micronuclei arising from chromosomes lagging at anaphase (Ogielska, 1994;Dedukh et al, 2019;Dedukh et al, 2022b). These small chromatin bodies contain the rejected chromosomes (Dedukh et al, 2017;Dedukh et al, 2019;Chmielewska et al, 2018;Dedukh et al, 2020b).…”