Olga Jablonska

Hybridization and polyploidization are key evolutionary forces shaping fish biodiversity. But their interaction with environmental factors, such as temperature, remains poorly understood. This study examined how maternal genome composition and incubation water temperature influence the hatching success, ploidy structure, morphology and early growth of polyploid Cobitis larvae. Crosses were performed using triploid Cobitis females with three genomic compositions (EEN, EET and ETN), representing combinations of genomes from C. elongatoides (E), C. taenia (T) and C. tanaitica (N), and diploid C. taenia males as sperm donors. Fertilized eggs were incubated at 18 °C, 22 °C and 28 °C. Triploid and tetraploid offspring occurred in comparable proportions on average across all groups, but developmental abnormalities were significantly more observed in tetraploid larvae. Females with EET and ETN genomes achieved higher hatching success than those with the EEN genome. Temperature had a pronounced effect on developmental timing and success: hatching occurred earliest at 28 °C, but survival decreased and abnormalities were most frequent. These results highlight genome- and temperature-dependent trade-offs in early development of Cobitis hybrids, providing new insight into reproductive dynamics and the potential resilience of polyploid systems under climate warming.

Toll-like receptors (TLRs) are key components of the innate immune system in fish, responsible for recognizing pathogen-associated molecular patterns derived from bacteria, viruses, and fungi. The sterlet (Acipenser ruthenus), an endangered sturgeon species valued for its meat and caviar, is a promising model for studying the effects of polyploidy on immune gene regulation. This study examined the expression of Toll-like receptor type 2 (TLR2) and type 13 (TLR13) in the heart, liver, gills, spleen, and kidney of diploid and triploid healthy sterlets using real-time PCR. TLR2 and TLR13 were expressed in all tissues of both diploids and triploids. In diploids, TLR2 expression was the highest in the kidney and the lowest in the liver (p < 0.05). Similarly, TLR13 expression in diploids was highest in the kidney and gills, and lowest in the liver (p < 0.05). In triploids, no significant tissue-specific variation in TLR expression was observed (p > 0.05). Comparisons between diploid and triploid sterlets revealed higher TLR2 expression in the kidney and higher TLR13 expression in the heart and kidney of diploids (p < 0.05). These molecular findings were supported by leukocyte analysis, which showed a significantly lower percentage of lymphocytes and a higher proportion of neutrophils in triploids compared to diploids. Additionally, the proportion of thrombocytes was significantly elevated in triploids (p < 0.05). This study provides the first report of TLR expression in polyploid fish, offering new insights into immune modulation associated with polyploidy in sturgeons.

Background: Interspecific hybridization between relative species Cobitis taenia (with a diploid genome designated as TT), Cobitis elongatoides (EE) and Cobitis tanaitica (NN) and the successive polyploidization with transitions from sexuality to asexuality experienced by triploid Cobitis hybrids likely influence their chromosomal rearrangements, including rearrangements of ribosomal DNA (rDNA) distribution patterns. Previously, we documented distinct karyotypic differences: C. elongatoides exhibited bi-armed chromosomes while C. taenia showed uni-armed chromosomes with rDNA-positive hybridization signals, respectively. Methods: In this study, fluorescence in situ hybridization (FISH) with 5S rDNA and 28S rDNA probes was used to analyze and compare chromosomal distribution patterns of rDNAs in clonally reproduced triploid Cobitis hybrids of different genomic constitutions ETT, ETN, EEN and EET (referred to using acronyms denoting the haploid genomes of their parent species), and their parental species. Results: Cobitis triploid hybrids exhibited intermediate karyotypes with ribosome synthesis sites on chromosomes inherited from both parents, showing no evidence of nucleolar dominance. The rDNA pattern derived from the C. elongatoides genome was more stable in the hybrids’ karyotypes. Two and one submetacentric chromosomes with co-localized rDNAs were effective markers to ascertain C. elongatoides diploid (EE) and haploid (E) genomes within the genome of triploid hybrids, respectively. Fewer 5S rDNA loci were observed in diploid (TT) and haploid (T) chromosome sets from C. taenia in ETT and EET females. C. taenia and C. tanaitica exhibited similar rDNA distribution patterns. Conclusions: The karyotypes of triploid Cobitis hybrids reflect the genomic contributions of their parental species. Variability in rDNA distribution patterns suggests complex genomic interactions in Cobitis hybrids resulting from polyploidization and hybridization, potentially influencing their reproductive potential.

Cobitis species exist in diploid populations, but mostly they occur in diploid-polyploid (d-p) ones. They are considered an important model organism to study biology and physiology of natural hybrid and polyploid vertebrates. Indeed, polyploidization causes a huge stress for the cell physiology and alter spermatogenesis in polyploid fish. The most extensively studied modes of germ cell death during spermatogenesis in vertebrates is apoptosis. The aim of the study was to examine the caspase-3 immunoexpression in the testes of Cobitis taenia from diploid population as well as C. taenia and sterile tetraploid Cobitis from d-p population before, during and after spawning. The obtained results suggest a different performance of apoptosis in testes of C. taenia from both studied populations and seems to be conditioned by their role as the only sperm donors in d-p populations. Moreover, apoptosis was active cell death process in the testes of tetraploid Cobitis.

Hybridization and polyploidization are important evolutionary processes whose impacts range from the alteration of gene expression and phenotypic variation to the triggering of asexual reproduction. We investigated fishes of the Cobitis taenia-elongatoides hybrid complex, which allowed us to disentangle the direct effects of both processes, due to the co-occurrence of parental species with their diploid and triploid hybrids. Employing morphological, ecological, and RNAseq approaches, we investigated the molecular determinants of hybrid and polyploid forms. In contrast with other studies, hybridization and polyploidy induced relatively very little transgressivity. Instead, Cobitis hybrids appeared intermediate with a clear effect of genomic dosing when triploids expressed higher similarity to the parent contributing two genome sets. This dosage effect was symmetric in the germline (oocyte gene expression), interestingly though, we observed an overall bias toward C. taenia in somatic tissues and traits. At the level of individual genes, expression-level dominance vastly prevailed over additivity or transgressivity. Also, trans-regulation of gene expression was less efficient in diploid hybrids than in triploids, where the expression modulation of homoeologs derived from the “haploid” parent was stronger than those derived from the “diploid” parent. Our findings suggest that the apparent intermediacy of hybrid phenotypes results from the combination of individual genes with dominant expression rather than from simple additivity. The efficiency of cross-talk between trans-regulatory elements further appears dosage dependent. Important effects of polyploidization may thus stem from changes in relative concentrations of trans-regulatory elements and their binding sites between hybridizing genomes. Links between gene regulation and asexuality are discussed.

The diploid–polyploid populations of Cobitis distributed in Poland are usually composed of the spined loach Cobitis taenia or, less often, the Danubian loach C. elongatoides and their triploid (females) and tetraploid hybrids (females and males). The aim of this study was to determine whether tetraploid males participate in the reproduction process by analyzing their testis ultrastructure and the process of spermatogenesis in comparison with diploid males of both parental species. Tetraploid loaches were obtained from three different diploid–polyploid populations distributed in Poland. The structure of Cobitis testes are typical for most Teleostei fish with cystic-type spermatogenesis. The successive stages of developing germ cells are enclosed within cysts formed by the Sertoli cells. This paper morphologically describes the different germ cell stages of spermatogenesis (spermatogonia, spermatocytes, spermatids, and spermatozoa) of C. taenia and C. elongatoides and provides a pioneering ultrastructural analysis of tetraploid Cobitis testes which reveals their unusual structure for the first time. Thus, cysts with normal spermatogonia and spermatocytes (pachyten or leptoten stages) containing synaptonemal complexes were present and no spermatids or spermatozoa were observed. Moreover, in contrast to previously analyzed diploid species, single cells or all of the cells within the cysts displayed chromatin condensation and/or chromatin fragmentation. The obtained results clearly demonstrated that tetraploid males are sterile and diploids are fertile and are the only sperm donors in the reproduction processes of diploid–polyploid Cobitis populations.

Fifteen microsatellite loci were identified in the tetraploid spined loach, Cobitis biwae (Teleostei: Cobitidae). Among these, 14 were polymorphic (5-31 alleles) and showed moderate to high cross-species amplification transferability in four related species, Cobitis matsubarai, Cobitis taenia, Misgurnus anguillicaudatus, and Misgurnus fossilis. The loci, described herein, will be useful for population genetics, phylogeny, parentage analysis, and detection of hybridization among Cobitis species.

Low doses of endocrine disrupting chemicals (EDCs) used in combination may act in a manner different from that of individual compounds. The objective of the study was to examine in vitro effects of low doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 100 pM) and genistein (500 nM) on: 1) progesterone (P4) and estradiol (E2) secretion (48 h); 2) dynamic changes in aryl hydrocarbon receptor (AhR) mRNA and protein expression (1, 3, 6, 24 and 48 h); 3) dynamic changes in estrogen receptor β (ERβ) mRNA and protein expression (1, 3, 6, 24 and 48 h); and 4) induction of apoptosis in porcine granulosa cells derived from medium follicles (3, 6 and 24 h). TCDD had no effect on P4 or E2 production, but potentiated the inhibitory effect of genistein on P4 production. In contrast to the individual treatments which did not produce any effects, TCDD and genistein administered together decreased ERβ and AhR protein expression in granulosa cells. Moreover, the inhibitory effect of TCDD on AhR mRNA expression was abolished by genistein. The treatments did not induce apoptosis in the cells. In summary, combined effects of low concentrations of TCDD and genistein on follicular function of pigs differed from that of individual compounds. The results presented in the current paper clearly indicate that effects exerted by low doses of EDCs applied in combination must be taken into consideration when studying potential risk effects of EDCs on biological processes.