Loci Associated with Negative Heterosis for Viability and Meat Productivity in Interspecific Sheep Hybrids.

Negative heterosis can occur on different economically important traits, but the exact biological mechanisms of this phenomenon are still unknown. The present study focuses on determining the genetic factors associated with negative heterosis in interspecific hybrids between domestic sheep (Ovis aries) and argali (Ovis ammon). One locus (rs417431015) associated with viability and two loci (rs413302370, rs402808951) associated with meat productivity were identified. One gene (ARAP2) was prioritized for viability and three for meat productivity (PDE2A, ARAP1, and PCDH15). The loci associated with meat productivity were demonstrated to fit the overdominant inheritance model and could potentially be involved int negative heterosis mechanisms.

Establishment of the Primary Avian Gonadal Somatic Cell Lines for Cytogenetic Studies.

The last decade was marked by a steep rise in avian studies at genomic and cellular levels. Cell lines are important tools for in vitro studies in cell biology and cytogenetics. We developed a simple method of primary somatic cell culture establishment from the ovaries of the great tits (Parus major) and testes of ten Passerine species, characterized the cellular composition of the ovary-derived lines using RT-PCR and immunolocalization of the tissue-specific markers and tested the efficiency of two methods of genetic transformation of the ovary-derived cell line. We found that the ovary-derived cell cultures of the great tit were composed of fibroblasts mainly, but also contained interstitial and granulosa cells. They were cultivated until the 10th passage without any noticeable decrease in their proliferative activity. The testis-derived cell cultures had lower proliferative potential. However, both ovary- and testis-derived cell cultures provided enough material for high quality mitotic metaphase chromosome preparations. The efficiency of its transduction with lentivirus containing a GFP reporter was very low, while electroporation with episomal vectors expressing GFP resulted in a high yield of GFP-positive cells. The proposed method could be used for the generation of high quality material for various cytogenetic and genomic studies.

Germline-Restricted Chromosomes and Autosomal Variants Revealed by Pachytene Karyotyping of 17 Avian Species.

Karyotypes of less than 10% of bird species are known. Using immunolocalization of the synaptonemal complex, the core structure of meiotic chromosomes at the pachytene stage, and centromere proteins, we describe male pachytene karyotypes of 17 species of birds. This method enables higher resolution than the conventional analyses of metaphase chromosomes. We provide the first descriptions of the karyotypes of 3 species (rook, Blyth's reed warbler, and European pied flycatcher), correct the published data on the karyotypes of 10 species, and confirm them for 4 species. All passerine species examined have highly conservative karyotypes, 2n = 80-82 with 7 pairs of macrochromosomes (including the ZZ sex chromosome pair which was not unambiguously distinguished from other macrochromosomes in most species) and 33-34 pairs of microchromosomes. In all of them, but not in the common cuckoo, we revealed single copies of the germline-restricted chromosomes varying in size and morphology even between closely related species. This indicates a fast evolution of this additional chromosome. The interspecies differences concern the sizes of the macrochromosomes, morphology of the microchromosomes, and sizes of the centromeres. The pachytene cells of the gouldian finch, brambling, and common linnet contain heteromorphic synaptonemal complexes indicating heterozygosity for inversions or centromere shifts. The European pied flycatcher, gouldian finch, and domestic canary have extended centromeres in several macro- and microchromosomes.

Whole-chromosome fusions in the karyotype evolution of Sceloporus (Iguania, Reptilia) are more frequent in sex chromosomes than autosomes.

Whole-chromosome fusions play a major role in the karyotypic evolution of reptiles. It has been suggested that certain chromosomes tend to fuse with sex chromosomes more frequently than others. However, the comparative genomic synteny data are too scarce to draw strong conclusions. We obtained and sequenced chromosome-specific DNA pools of Sceloporus malachiticus, an iguanian species which has experienced many chromosome fusions. We found that four of seven lineage-specific fusions involved sex chromosomes, and that certain syntenic blocks which constitute the sex chromosomes, such as the homologues of the Anolis carolinensis chromosomes 11 and 16, are repeatedly involved in sex chromosome formation in different squamate species. To test the hypothesis that the karyotypic shift could be associated with changes in recombination patterns, we performed a synaptonemal complex analysis in this species and in Sceloporus variabilis (2n = 34). It revealed that the sex chromosomes in S. malachiticus had two distal pseudoautosomal regions and a medial differentiated region. We found that multiple fusions little affected the recombination rate in S. malachiticus. Our data confirm more frequent involvement of certain chromosomes in sex chromosome formation, but do not reveal a connection between the gonosome-autosome fusions and the evolution of recombination rate. This article is part of the theme issue 'Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part II)'.

Reproductive Isolation Between Taxonomically Controversial Forms of the Gray Voles (Microtus, Rodentia; Arvicolinae): Cytological Mechanisms and Taxonomical Implications.

The formation of hybrid sterility is an important stage of speciation. The voles of the genus Microtus, which is the most speciose genus of rodents, provide a good model for studying the cytological mechanisms of hybrid sterility. The voles of the "mystacinus" group of the subgenus Microtus (2n = 54) comprising several recently diverged forms with unclear taxonomic status are especially interesting. To resolve the taxonomic status of Microtus mystacinus and Microtus kermanensis, we crossed both with Microtus rossiaemeridionalis, and M. kermanensis alone with Microtus arvalis "obscurus" and M. transcaspicus and examined the reproductive performance of their F1 hybrids. All interspecies male hybrids were sterile. Female M. kermanensis × M. arvalis and M. kermanensis × M. transcaspicus hybrids were sterile as well. Therefore, M. mystacinus, M. kermanensis, and M. rossiaemeridionalis could be considered valid species. To gain an insight into the cytological mechanisms of male hybrid sterility, we carried out a histological analysis of spermatogenesis and a cytological analysis of chromosome synapsis, recombination, and epigenetic chromatin modifications in the germ cells of the hybrids using immunolocalization of key meiotic proteins. The hybrids showed wide variation in the onset of spermatogenesis arrest stage, from mature (although abnormal) spermatozoa to spermatogonia only. Chromosome asynapsis was apparently the main cause of meiotic arrest. The degree of asynapsis varied widely across cells, individuals, and the crosses-from partial asynapsis of several small bivalents to complete asynapsis of all chromosomes. The asynapsis was accompanied by a delayed repair of DNA double-strand breaks marked by RAD51 antibodies and silencing of unpaired chromatin marked by γH2A.X antibodies. Overall, the severity of disturbances in spermatogenesis in general and in chromosome synapsis in particular increased in the hybrids with an increase in the phylogenetic distance between their parental species.

Multivariate Analysis Identifies Eight Novel Loci Associated with Meat Productivity Traits in Sheep.

Despite their economic value, sheep remain relatively poorly studied animals in terms of the number of known loci and genes associated with commercially important traits. This gap in our knowledge can be filled in by performing new genome-wide association studies (GWAS) or by re-analyzing previously documented data using novel powerful statistical methods. This study is focused on the search for new loci associated with meat productivity and carcass traits in sheep. With a multivariate approach applied to publicly available GWAS results, we identified eight novel loci associated with the meat productivity and carcass traits in sheep. Using an in silico follow-up approach, we prioritized 13 genes in these loci. One of eight novel loci near the FAM3C and WNT16 genes has been replicated in an independent sample of Russian sheep populations (N = 108). The novel loci were added to our regularly updated database increasing the number of known loci to more than 140.

Homogeneously Staining Regions (HSR) in Chromosome 1 of the House Mouse: Synapsis and Recombination at Meiosis.

Amplified sequences constitute a large part of mammalian genomes. A chromosome 1 containing 2 large (up to 50 Mb) homogeneously staining regions (HSRs) separated by a small inverted euchromatic region is present in many natural populations of the house mouse (Mus musculus musculus). The HSRs are composed of a long-range repeat cluster, Sp100-rs, with a repeat length of 100 kb. In order to understand the organization and function of HSRs in meiotic chromosomes, we examined synapsis and recombination in male mice hetero- and homozygous for the HSR-carrying chromosome using FISH with an HSR-specific DNA probe and immunolocalization of the key meiotic proteins. In all homozygous and heterozygous pachytene nuclei, we observed fully synapsed linear homomorphic bivalents 1 marked by the HSR FISH probe. The synaptic adjustment in the heterozygotes was bilateral: the HSR-carrying homolog was shortened and the wild-type homolog was elongated. The adjustment was reversible: desynapsis at diplotene was accompanied by elongation of the HSRs. Immunolocalization of H3K9me2/3 indicated that the HSRs in the meiotic chromosome retained the epigenetic modification typical for C-heterochromatin in somatic cells. MLH1 foci, marking mature recombination nodules, were detected in the proximal HSR band in heterozygotes and in both HSR bands of homozygotes. Unequal crossing over within the long-range repeat cluster can cause variation in size of the HSRs, which has been detected in the natural populations of the house mouse.

Heterochiasmy and Sexual Dimorphism: The Case of the Barn Swallow (Hirundo rustica, Hirundinidae, Aves).

Heterochiasmy, a sex-based difference in recombination rate, has been detected in many species of animals and plants. Several hypotheses about evolutionary causes of heterochiasmy were proposed. However, there is a shortage of empirical data. In this paper, we compared recombination related traits in females and males of the barn swallow Hirundo rustica (Linnaeus, 1758), the species under strong sexual selection, with those in the pale martin Riparia diluta (Sharpe and Wyatt, 1893), a related and ecologically similar species with the same karyotype (2N = 78), but without obvious sexual dimorphism. Recombination traits were examined in pachytene chromosome spreads prepared from spermatocytes and oocytes. Synaptonemal complexes and mature recombination nodules were visualized with antibodies to SYCP3 and MLH1 proteins, correspondingly. Recombination rate was significantly higher (p = 0.0001) in barn swallow females (55.6 ± 6.3 recombination nodules per autosomal genome), caused by the higher number of nodules at the macrochromosomes, than in males (49.0 ± 4.5). They also showed more even distribution of recombination nodules along the macrochromosomes. At the same time, in the pale martin, sexual differences in recombination rate and distributions were rather small. We speculate that an elevated recombination rate in the female barn swallows might have evolved as a compensatory reaction to runaway sexual selection in males.

Chromosome Painting Does Not Support a Sex Chromosome Turnover in Lacerta agilis Linnaeus, 1758.

Reptiles show a remarkable diversity of sex determination mechanisms and sex chromosome systems, derived from different autosomal pairs. The origin of the ZW sex chromosomes of Lacerta agilis, a widespread Eurasian lizard species, is a matter of discussion: is it a small macrochromosome from the 11-18 group common to all lacertids, or does this species have a unique ZW pair derived from the large chromosome 5? Using independent molecular cytogenetic methods, we investigated the karyotype of L. agilis exigua from Siberia, Russia, to identify the sex chromosomes. FISH with a flow-sorted chromosome painting probe derived from L. strigata and specific to chromosomes 13, 14, and Z confirmed that the Z chromosome of L. agilis is a small macrochromosome, the same as in L. strigata. FISH with the telomeric probe showed an extensive accumulation of the telomere-like repeat in the W chromosome in agreement with previous studies, excluding the possibility that the lineages of L. agilis studied in different works could have different sex chromosome systems due to a putative intra-species polymorphism. Our results reinforce the idea of the stability of the sex chromosomes and lack of evidence for sex-chromosome turnovers in known species of Lacertidae.

Germline-restricted chromosome (GRC) in the sand martin and the pale martin (Hirundinidae, Aves): synapsis, recombination and copy number variation.

All songbirds studied to date have an additional Germline Restricted Chromosome (GRC), which is not present in somatic cells. GRCs show a wide variation in genetic content and little homology between species. To check how this divergence affected the meiotic behavior of the GRC, we examined synapsis, recombination and copy number variation for GRCs in the closely related sand and pale martins (Riparia riparia and R. diluta) in comparison with distantly related estrildid finches. Using immunolocalization of meiotic proteins and FISH with GRC-specific DNA probes, we found a striking similarity in the meiotic behavior of GRCs between martins and estrildid finches despite the millions of years of independent evolution. GRCs are usually present in two copies in female and in one copy in male pachytene cells. However, we detected polymorphism in female and mosaicism in male martins for the number of GRCs. In martin and zebra finch females, two GRCs synapse along their whole length, but recombine predominately at their ends. We suggest that the shared features of the meiotic behavior of GRCs have been supported by natural selection in favor of a preferential segregation of GRCs to the eggs.

Interbreed variation in meiotic recombination rate and distribution in the domestic chicken Gallus gallus.

The efficiency of natural and artificial selection is critically dependent on the recombination rate. However, interbreed and individual variation in recombination rate in poultry remains unknown. Conventional methods of analysis of recombination such as genetic linkage analysis, sperm genotyping and chiasma count at lampbrush chromosomes are expensive and time-consuming. In this study, we analyzed the number and distribution of recombination nodules in spermatocytes of the roosters of six chicken breeds using immunolocalization of key proteins involved in chromosome pairing and recombination. We revealed significant effects of breed ( R 2 = 0.17 ; p < 0.001 ) and individual ( R 2 = 0.28 ; p < 0.001 ) on variation in the number of recombination nodules. Both interbreed and individual variations in recombination rate were almost entirely determined by variation in recombination density on macrochromosomes, because almost all microchromosomes in each breed had one recombination nodule. Despite interbreed differences in the density of recombination nodules, the patterns of their distribution along homologous chromosomes were similar. The breeds examined in this study showed a correspondence between the age of the breed and its recombination rate. Those with high recombination rates (Pervomai, Russian White and Brahma) are relatively young breeds created by crossing several local breeds. The breeds displaying low recombination rate are ancient local breeds: Cochin (Indo-China), Brown Leghorn (Tuscany, Italy) and Russian Crested (the European part of Russia).

Identification of sex chromosomes in Eremiasvelox (Lacertidae, Reptilia) using lampbrush chromosome analysis.

Reptiles are good objects for studying the evolution of sex determination, since they have different sex determination systems in different lineages. Lacertid lizards have been long-known for possessing ZZ/ZW type sex chromosomes. However, due to morphological uniformity of lacertid chromosomes, the Z chromosome has been only putatively cytologically identified. We used lampbrush chromosome (LBC) analysis and FISH with a W-specific probe in Eremiasvelox (Pallas, 1771) to unequivocally identify the ZW bivalent and investigate its meiotic behavior. The heterochromatic W chromosome is decondensed at the lampbrush stage, indicating active transcription, contrast with the highly condensed condition of the lampbrush W chromosomes in birds. We identified the Z chromosome by its chiasmatic association with the W chromosome as chromosome XIII of the 19 chromosomes in the LBC karyotype. Our findings agree with previous genetic and genomic studies, which suggested that the lacertid Z chromosome should be one of the smaller macrochromosomes.

Germline-restricted chromosome (GRC) is widespread among songbirds.

An unusual supernumerary chromosome has been reported for two related avian species, the zebra and Bengalese finches. This large, germline-restricted chromosome (GRC) is eliminated from somatic cells and spermatids and transmitted via oocytes only. Its origin, distribution among avian lineages, and function were mostly unknown so far. Using immunolocalization of key meiotic proteins, we found that GRCs of varying size and genetic content are present in all 16 songbird species investigated and absent from germline genomes of all eight examined bird species from other avian orders. Results of fluorescent in situ hybridization of microdissected GRC probes and their sequencing indicate that GRCs show little homology between songbird species and contain a variety of repetitive elements and unique sequences with paralogs in the somatic genome. Our data suggest that the GRC evolved in the common ancestor of all songbirds and underwent significant changes in the extant descendant lineages.

Male Meiotic Recombination in the Steppe Agama, Trapelus sanguinolentus (Agamidae, Iguania, Reptilia).

Meiotic recombination rates and patterns of crossover distributions along the chromosomes vary considerably even between closely related species. The adaptive significance of these differences is still unclear due to the paucity of empirical data. Most data on recombination come from mammalian species, while other vertebrate clades are poorly explored. Using immunolocalization of the protein of the lateral element of the synaptonemal complex (SYCP3) and the mismatch-repair protein MLH1, which marks mature recombination nodules, we analyzed recombination rates and crossover distribution in meiotic prophase chromosomes of the steppe agama (Trapelus sanguinolentus, Agamidae, Acrodonta, Iguania) and compared them with data obtained for the genus Anolis (Dactyloidae, Pleurodonta, Iguania). We found that, despite a smaller genome size, the total SC length and the MLH1 focus number per cell are much higher in the agama than in the anoles. The distributions of the MLH1 foci in the agama are multimodal in larger chromosomes and bimodal in smaller chromosomes without a significant centromere effect, resembling the patterns known for birds. A possible relationship between karyotype remodeling and the evolution of recombination in Iguania is discussed.

Recent advances in understanding genetic variants associated with growth, carcass and meat productivity traits in sheep (Ovis aries): an update.

Identification of quantitative trait loci (QTLs) and candidate genes that affect growth intensity is a prerequisite for the marker-assisted selection of economically important traits. The number of QTL studies on sheep is relatively small in comparison to those on cattle and pigs. The current QTL sheep database - Sheep QTLdb - contains information on 1658 QTLs for 225 different traits. A few genes and markers associated with growth, carcass and meat productivity traits have been reported. The information about QTLs from the Sheep QTLdb cannot be directly used in marker-assisted selection due to the lack of essential information such as effective and reference alleles, the effect direction etc., and it requires manual curation and validation. In this study we performed a comprehensive search for QTLs focusing on single nucleotide polymorphisms (SNPs) associated with growth and meat traits in sheep. The database contains information about 156 SNP-trait associations (123 unique SNPs) and a list of 165 associated genes. The updated information is freely available at https://github.com/Defrag1236/Ovines_2018 (last access: 18 September 2019). This information can be useful for further association studies and preliminary estimation of genetic variability for economically important traits in different breeds.

Chromosome Synapsis and Recombination in Male-Sterile and Female-Fertile Interspecies Hybrids of the Dwarf Hamsters (Phodopus, Cricetidae).

Hybrid sterility is an important step in the speciation process. Hybrids between dwarf hamsters Phodopus sungorus and P.campbelli provide a good model for studies in cytological and genetic mechanisms of hybrid sterility. Previous studies in hybrids detected multiple abnormalities of spermatogenesis and a high frequency of dissociation between the X and Y chromosomes at the meiotic prophase. In this study, we found that the autosomes of the hybrid males and females underwent paring and recombination as normally as their parental forms did. The male hybrids showed a significantly higher frequency of asynapsis and recombination failure between the heterochromatic arms of the X and Y chromosomes than the males of the parental species. Female hybrids as well as the females of the parental species demonstrated a high incidence of centromere misalignment at the XX bivalent and partial asynapsis of the ends of its heterochromatic arms. In all three karyotypes, recombination was completely suppressed in the heterochromatic arm of the X chromosome, where the pseudoautosomal region is located. We propose that this recombination pattern speeds up divergence of the X- and Y-linked pseudoautosomal regions between the parental species and results in their incompatibility in the male hybrids.

Chromosome Synapsis and Recombination in Male Hybrids between Two Chromosome Races of the Common Shrew (Sorex araneus L., Soricidae, Eulipotyphla).

Hybrid zones between chromosome races of the common shrew (Sorex araneus) provide exceptional models to study the potential role of chromosome rearrangements in the initial steps of speciation. The Novosibirsk and Tomsk races differ by a series of Robertsonian fusions with monobrachial homology. They form a narrow hybrid zone and generate hybrids with both simple (chain of three chromosomes) and complex (chain of eight or nine) synaptic configurations. Using immunolocalisation of the meiotic proteins, we examined chromosome pairing and recombination in males from the hybrid zone. Homozygotes and simple heterozygotes for Robertsonian fusions showed a low frequency of synaptic aberrations (<10%). The carriers of complex synaptic configurations showed multiple pairing abnormalities, which might lead to reduced fertility. The recombination frequency in the proximal regions of most chromosomes of all karyotypes was much lower than in the other regions. The strong suppression of recombination in the pericentromeric regions and co-segregation of race specific chromosomes involved in the long chains would be expected to lead to linkage disequilibrium between genes located there. Genic differentiation, together with the high frequency of pairing aberrations in male carriers of the long chains, might contribute to maintenance of the narrow hybrid zone.

Immunocytological analysis of meiotic recombination in two anole lizards (Squamata, Dactyloidae).

Although the evolutionary importance of meiotic recombination is not disputed, the significance of interspecies differences in the recombination rates and recombination landscapes remains under-appreciated. Recombination rates and distribution of chiasmata have been examined cytologically in many mammalian species, whereas data on other vertebrates are scarce. Immunolocalization of the protein of the synaptonemal complex (SYCP3), centromere proteins and the mismatch-repair protein MLH1 was used, which is associated with the most common type of recombination nodules, to analyze the pattern of meiotic recombination in the male of two species of iguanian lizards, Anolis carolinensis Voigt, 1832 and Deiroptyx coelestinus (Cope, 1862). These species are separated by a relatively long evolutionary history although they retain the ancestral iguanian karyotype. In both species similar and extremely uneven distributions of MLH1 foci along the macrochromosome bivalents were detected: approximately 90% of crossovers were located at the distal 20% of the chromosome arm length. Almost total suppression of recombination in the intermediate and proximal regions of the chromosome arms contradicts the hypothesis that "homogenous recombination" is responsible for the low variation in GC content across the anole genome. It also leads to strong linkage disequilibrium between the genes located in these regions, which may benefit conservation of co-adaptive gene arrays responsible for the ecological adaptations of the anoles.

Spatial organization of fibroblast and spermatocyte nuclei with different B-chromosome content in Korean field mouse, Apodemus peninsulae (Rodentia, Muridae).

Korean field mouse (Apodemus peninsulae) shows a wide variation in the number of B chromosomes composed of constitutive heterochromatin. For this reason, it provides a good model to study the influence of the number of centromeres and amount of heterochromatin on spatial organization of interphase nuclei. We analyzed the three-dimensional organization of fibroblast and spermatocyte nuclei of the field mice carrying a different number of B chromosomes using laser scanning microscopy and 3D fluorescence in situ hybridization. We detected a co-localization of the B chromosomes with constitutive heterochromatin of the chromosomes of the basic set. We showed a non-random distribution of B chromosomes in the spermatocyte nuclei. Unpaired B chromosomes showed a tendency to occur in the compartment formed by the unpaired part of the XY bivalent.