Hidden Compositional Heterogeneity of Fish Chromosomes in the Era of Polished Genome Assemblies

Fish chromosomes are considered homogeneous in their AT/GC nucleotide composition, and banding patterns enabling identification of homologs are largely missing. While cytogenomic approaches try to compensate for this issue by virtual karyotyping, they rely on the quality of genome assemblies available. Recently, soft-masked genome assemblies combining costly and arduous long- and short-read sequencing and new generation assemblers became available for two teleost fish species, climbing perch (Anabas testudineus) and channel bull blenny (Cottoperca gobio). Soft-masking turns repetitive sequences in a genome assembly into lower case letters, leaving unique sequences in upper case. This enables investigators to assess the proportion of guanine and cytosine nucleotides (GC%) of transposable elements as an indicator of AT/GC homogenisation in fish. We have developed a new version of our Python tool Evan, which utilises chromosome-level genome assemblies and combines the profiles of GC% and the proportion of repeats (rep%) along chromosomes. Our profiles of both of those fishes showed clear and abrupt but small-scale fluctuations in GC% along otherwise compositionally homogenised sequences. Our study also highlights the key role of the sliding window size in determining the resolution of GC% profiling. While the quality of the genome assemblies appeared to be sufficient for GC%/rep% profiling, more effective repeat masking is necessary to better distinguish to what extent repeats compositionally homogenize fish genomes.journal articl

The second highest chromosome count among vertebrates is observed in cultured sturgeon and is associated with genome plasticity

Additional file 1: Table S1. Genotyping results with highlighted private dam and sire microsatellite alleles observed at all analysed loci for parental individuals, and for pentaploid (5n) and heptaploid (7n) full siblings

Karyotype differentiation in 19 species of river loach fishes (Nemacheilidae, Teleostei): extensive variability associated with rDNA and heterochromatin distribution and its phylogenetic and ecological interpretation

GenBank accession numbers of cyt b, IRBP and RAG1 sequences of nemacheilids and one botiid species (B. lohachata). (XLSX 17 kb

Molecular cytogenetic differentiation of paralogs of Hox paralogs in duplicated and re-diploidized genome of the North American paddlefish (Polyodon spathula).

BackgroundAcipenseriformes is a basal lineage of ray-finned fishes and comprise 27 extant species of sturgeons and paddlefishes. They are characterized by several specific genomic features as broad ploidy variation, high chromosome numbers, presence of numerous microchromosomes and propensity to interspecific hybridization. The presumed palaeotetraploidy of the American paddlefish was recently validated by molecular phylogeny and Hox genes analyses. A whole genome duplication in the paddlefish lineage was estimated at approximately 42 Mya and was found to be independent from several genome duplications evidenced in its sister lineage, i.e. sturgeons. We tested the ploidy status of available chromosomal markers after the expected rediploidization. Further we tested, whether paralogs of Hox gene clusters originated from this paddlefish specific genome duplication are cytogenetically distinguishable.ResultsWe found that both paralogs HoxA alpha and beta were distinguishable without any overlapping of the hybridization signal - each on one pair of large metacentric chromosomes. Of the HoxD, only the beta paralog was unequivocally identified, whereas the alpha paralog did not work and yielded only an inconclusive diffuse signal. Chromosomal markers on three diverse ploidy levels reflecting different stages of rediploidization were identified: quadruplets retaining their ancestral tetraploid condition, semi-quadruplets still reflecting the ancestral tetraploidy with clear signs of advanced rediploidization, doublets were diploidized with ancestral tetraploidy already blurred. Also some of the available microsatellite data exhibited diploid allelic band patterns at their loci whereas another locus showed more than two alleles.ConclusionsOur exhaustive staining of paddlefish chromosomes combined with cytogenetic mapping of ribosomal genes and Hox paralogs and with microsatellite data, brings a closer look at results of the process of rediploidization in the course of paddlefish genome evolution. We show a partial rediploidization represented by a complex mosaic structure comparable with segmental paleotetraploidy revealed in sturgeons (Acipenseridae). Sturgeons and paddlefishes with their high propensity for whole genome duplication thus offer suitable animal model systems to further explore evolutionary processes that were shaping the early evolution of all vertebrates

Higher-order organisation of extremely amplified, potentially functional and massively methylated 5S rDNA in European pikes (Esox sp.)

Background Pikes represent an important genus (Esox) harbouring a pre-duplication karyotype (2n = 2x = 50) of economically important salmonid pseudopolyploids. Here, we have characterized the 5S ribosomal RNA genes (rDNA) in Esox lucius and its closely related E. cisalpinus using cytogenetic, molecular and genomic approaches. Intragenomic homogeneity and copy number estimation was carried out using Illumina reads. The higher-order structure of rDNA arrays was investigated by the analysis of long PacBio reads. Position of loci on chromosomes was determined by FISH. DNA methylation was analysed by methylation-sensitive restriction enzymes. Results The 5S rDNA loci occupy exclusively (peri)centromeric regions on 30-38 acrocentric chromosomes in both E. lucius and E. cisalpinus. The large number of loci is accompanied by extreme amplification of genes (>20,000 copies), which is to the best of our knowledge one of the highest copy number of rRNA genes in animals ever reported. Conserved secondary structures of predicted 5S rRNAs indicate that most of the amplified genes are potentially functional. Only few SNPs were found in genic regions indicating their high homogeneity while intergenic spacers were more heterogeneous and several families were identified. Analysis of 10-30 kb-long molecules sequenced by the PacBio technology (containing about 40% of total 5S rDNA) revealed that the vast majority (96%) of genes are organised in large several kilobase-long blocks. Dispersed genes or short tandems were less common (4%). The adjacent 5S blocks were directly linked, separated by intervening DNA and even inverted. The 5S units differing in the intergenic spacers formed both homogeneous and heterogeneous (mixed) blocks indicating variable degree of homogenisation between the loci. Both E. lucius and E. cisalpinus 5S rDNA was heavily methylated at CG dinucleotides. Conclusions Extreme amplification of 5S rRNA genes in the Esox genome occurred in the absence of significant pseudogenisation suggesting its recent origin and/or intensive homogenisation processes. The dense methylation of units indicates that powerful epigenetic mechanisms have evolved in this group of fish to silence amplified genes. We discuss how the higher-order repeat structures impact on homogenisation of 5S rDNA in the genome

Present and Future Salmonid Cytogenetics

Salmonids are extremely important economically and scientifically; therefore, dynamic developments in their research have occurred and will continue occurring in the future. At the same time, their complex phylogeny and taxonomy are challenging for traditional approaches in research. Here, we first provide discoveries regarding the hitherto completely unknown cytogenetic characteristics of the Anatolian endemic flathead trout, Salmo platycephalus, and summarize the presently known, albeit highly complicated, situation in the genus Salmo. Secondly, by outlining future directions of salmonid cytogenomics, we have produced a prototypical virtual karyotype of Salmo trutta, the closest relative of S. platycephalus. This production is now possible thanks to the high-quality genome assembled to the chromosome level in S. trutta via soft-masking, including a direct labelling of repetitive sequences along the chromosome sequence. Repetitive sequences were crucial for traditional fish cytogenetics and hence should also be utilized in fish cytogenomics. As such virtual karyotypes become increasingly available in the very near future, it is necessary to integrate both present and future approaches to maximize their respective benefits. Finally, we show how the presumably repetitive sequences in salmonids can change the understanding of the overall relationship between genome size and G+C content, creating another outstanding question in salmonid cytogenomics waiting to be resolved

Microanatomy and cytogenetics of non-marine ostracods-an insight into evolutionary biology of their reproductive modes

SUMMARYOF THEPh.D.THESIS The main objectivesof this dissertationwere to contributeto generarunderstandingand to fill some major gaps in our knowledgeof ostracodbiology.Ostracodsareotherwisea very well exploredgroup of crusta-cea :f ff:#'1"'';i,:T*"ortneirooosfil;I"TilffJ:,X'H::':;:: reconstructions,tn,..Lll?,1',i#r:T::il ff: "t"" usedinpataeoecorogicar knownabouthepatopancreasandnothins "oor,,Jl1ll,f"filH, ]i:.ffil:Further,there is no properdescriptionof the com ;':il'i:?l,Tll;l':*'*'ornsemnaton*":fflilli::fi:ff H*::: onryavairabledata on ,lxisted concerningthe karyologyof freshwaterostracods.rhe with exceptionssome 'eshwater ostracodkaryologycamefrom the i930s to 1950s *#jli:.:.,:ru$"::lt:_,:ri::1,i:iJ.["::.13[*:Nonetheress,tn"r"n,nn,lTi"""t"t"iffil]ogenetic remaleshadbeen*,*o "*.contextortheirreproduJtivemodesremaine;-;il::*o:nfoTl.."l';n";;u"''",n"For these reasons, ostracod microanatomyand cytogeneticsin rerationto theevotutionarybiologyof theirreproductiu",o0". *"of this thesis.Moreouer,other topicsreratedto ,n",'t" 'nu"tt'n"ted in the framework also taken into account. -r;J;._:::,:: tn"'t reproductionbiorogyhad been manipu,atorsinoucinsparililj:"gnJfl:ffi:nTffi:i1"J:fi#["il:obtainedby microscopicanatysisof the FRO in liduring holotomographic Investigation of cretac tng animals has been utilized..