Chromosomal and genomic analysis suggests single origin and high molecular differentiation of the B chromosome of Abracris flavolineata.

Supernumerary chromosomes (B chromosomes) have been an intriguing subject of study. Our understanding of the molecular differentiation of B chromosomes from an interpopulation perspective remains limited, with most analyses involving chromosome banding and mapping of a few sequences. To gain insights into the molecular composition, origin, and evolution of B chromosomes, we conducted cytogenetic and next-generation sequencing analysis of the repeatome in the grasshopper Abracris flavolineata across various populations. Our results unveiled the presence of B chromosomes in two newly investigated populations and described new satellite DNA sequences. While we observed some degree of genetic connection among A. flavolineata populations, our comparative analysis of genomes with and without B chromosomes provided evidence of two new B chromosome variants. These variants exhibited distinct compositions of various repeat classes, including transposable elements and satellite DNAs. Based on shared repeats, their chromosomal location, and the C-positive heterochromatin content on the B chromosome, these variants likely share a common origin but have undergone distinct molecular differentiation processes, resulting in varying degrees of heterochromatinization. Our data serve as a detailed example of the dynamic and differentiated nature of B chromosome molecular content at the interpopulation level, even when they share a common origin.

Evolution of satDNAs on holocentric chromosomes: insights from hemipteran insects of the genus Mahanarva.

Satellite DNAs (satDNAs) constitute one of the main components of eukaryote genomes and are involved in chromosomal organization and diversification. Although largely studied, little information was gathered about their evolution on holocentric species, i.e., diffuse centromeres, which, due to differences in repeat organization, could result in different evolutionary patterns. Here, we combined bioinformatics and cytogenetic approaches to evaluate the evolution of the satellitomes in Mahanarva holocentric insects. In two species, de novo identification revealed a high number of satDNAs, 110 and 113, with an extreme monomer length range of 18-4228 bp. The overall abundance of satDNAs was observed to be 6.67% in M. quadripunctata and 1.98% in M. spectabilis, with different abundances for the shared satDNAs. Chromosomal mapping of the most abundant repeats of M. quadripunctata and M. spectabilis on other Mahanarva reinforced the dynamic nature of satDNAs. Variable patterns of chromosomal distribution for the satDNAs were noticed, with the occurrence of clusters on distinct numbers of chromosomes and at different positions and the occurrence of scattered signals or nonclustered satDNAs. Altogether, our data demonstrated the high dynamism of satDNAs in Mahanarva with the involvement of this genomic fraction in chromosome diversification of the genus. The general characteristics and patterns of evolution of satDNAs are similar to those observed on monocentric chromosomes, suggesting that the differential organization of genome compartments observed on holocentric chromosomes compared with monocentric chromosomes does not have a large impact on the evolution of satDNAs. Analysis of the satellitomes of other holocentric species in a comparative manner will shed light on this issue.

A step forward in the genome characterization of the sugarcane borer, Diatraea saccharalis: karyotype analysis, sex chromosome system and repetitive DNAs through a cytogenomic approach.

Moths of the family Crambidae include a number of pests that cause economic losses to agricultural crops. Despite their economic importance, little is known about their genome architecture and chromosome evolution. Here, we characterized the chromosomes and repetitive DNA of the sugarcane borer Diatraea saccharalis using a combination of low-pass genome sequencing, bioinformatics, and cytogenetic methods, focusing on the sex chromosomes. Diploid chromosome numbers differed between the sexes, i.e., 2n = 33 in females and 2n = 34 in males. This difference was caused by the occurrence of a WZ1Z2 trivalent in female meiosis, indicating a multiple sex-chromosome system WZ1Z2/Z1Z1Z2Z2. A strong interstitial telomeric signal was observed on the W chromosome, indicating a fusion of the ancestral W chromosome with an autosome. Among repetitive DNAs, transposable elements (TEs) accounted for 39.18% (males) to 41.35% (females), while satDNAs accounted for only 0.214% (males) and 0.215% (females) of the genome. FISH mapping revealed different chromosomal organization of satDNAs, such as single localized clusters, spread repeats, and non-clustered repeats. Two TEs mapped by FISH were scattered. Although we found a slight enrichment of some satDNAs in the female genome, they were not differentially enriched on the W chromosome. However, we found enriched FISH signals for TEs on the W chromosome, suggesting their involvement in W chromosome degeneration and differentiation. These data shed light on karyotype and repetitive DNA dynamics due to multiple chromosome fusions in D. saccharalis, contribute to the understanding of genome structure in Lepidoptera and are important for future genomic studies.

The spread of satellite DNAs in euchromatin and insights into the multiple sex chromosome evolution in Hemiptera revealed by repeatome analysis of the bug Oxycarenus hyalinipennis.

Satellite DNAs (satDNAs) are highly repeated tandem sequences primarily located in heterochromatin, although their occurrence in euchromatin has been reported. Here, our aim was to advance the understanding of satDNA and multiple sex chromosome evolution in heteropterans. We combined cytogenetic and genomic approaches to study, for the first time, the satDNA composition of the genome in an Oxycarenidae bug, Oxycarenus hyalinipennis. The species exhibits a male karyotype of 2n = 19 (14A + 2 m + X1 X2 Y), with a highly differentiated Y chromosome, as demonstrated by C-banding and comparative genomic hybridization, revealing an enrichment of repeats from the male genome. Additionally, comparative analysis between males and females revealed that the 26 identified satDNA families are significantly biased towards male genome, accumulating in discrete regions in the Y chromosome. Exceptionally, the OhyaSat04-125 family was found to be distributed virtually throughout the entire extension of the Y chromosome. This suggests an important role of satDNA in Y chromosome differentiation, in comparison of other repeats, which collectively shows similar abundance between sexes, about 50%. Furthermore, chromosomal mapping of all satDNA families revealed an unexpected high spread in euchromatic regions, covering the entire extension, irrespective of their abundance. Only discrete regions of heterochromatin on the Y chromosome and of the m-chromosomes (peculiar chromosomes commonly observed in heteropterans) were enriched with satDNAs. The putative causes of the intense enrichment of satDNAs in euchromatin are discussed, including the possible existence of burst cycles similar to transposable elements and as a result of holocentricity. These data challenge the classical notion that euchromatin is not enriched with satDNAs.

The extensive amplification of heterochromatin in Melipona bees revealed by high throughput genomic and chromosomal analysis.

Satellite DNAs (satDNAs) and transposable elements (TEs) are among the main components of constitutive heterochromatin (c-heterochromatin) and are related to their functionality, dynamics, and evolution. A peculiar case regarding the quantity and distribution of c-heterochromatin is observed in the genus of bees, Melipona, with species having a low amount of heterochromatin and species with high amount occupying almost all chromosomes. By combining low-pass genome sequencing and chromosomal analysis, we characterized the satDNAs and TEs of Melipona quadrifasciata (low c-heterochromatin) and Melipona scutellaris (high low c-heterochromatin) to understand c-heterochromatin composition and evolution. We identified 15 satDNA families and 20 TEs for both species. Significant variations in the repeat landscapes were observed between the species. In M. quadrifasciata, the repetitive fraction corresponded to only 3.78% of the genome library studied, whereas in M. scutellaris, it represented 54.95%. Massive quantitative and qualitative changes contributed to the differential amplification of c-heterochromatin, mainly due to the amplification of exclusive repetitions in M. scutellaris, as the satDNA MscuSat01-195 and the TE LTR/Gypsy_1 that represent 38.20 and 14.4% of its genome, respectively. The amplification of these two repeats is evident at the chromosomal level, with observation of their occurrence on most c-heterochromatin. Moreover, we detected repeats shared between species, revealing that they experienced mainly quantitative variations and varied in the organization on chromosomes and evolutionary patterns. Together, our data allow the discussion of patterns of evolution of repetitive DNAs and c-heterochromatin that occurred in a short period of time, after separation of the Michmelia and Melipona subgenera.

The relevance of chromosome fissions for major ribosomal DNA dispersion in hymenopteran insects.

Ribosomal DNA (rDNA) loci are essential for cellular metabolism due to their participation in ribosome biogenesis. Although these genes have been widely cytogenetically mapped, the evolutionary mechanisms behind their variability in number and chromosomal location remain elusive, even in well-known biological groups, such as ants, bees and wasps (Insecta: Hymenoptera). To address this question in Hymenoptera and therefore advance the understanding of rDNA evolution in insects in general, we integrated molecular cytogenetic data, a phylogenomic framework, model-based predictions and genome sequencing. Hence, we assessed the main evolutionary trends shaping the chromosomal distribution of rDNA loci in Hymenoptera. We noticed the conservation of one site of rDNA per haploid genome, suggesting that a single 45S rDNA locus is the putative ancestral pattern for aculeate Hymenoptera. Moreover, our results highlighted a nonrandom distribution of rDNA in Hymenoptera karyotypes, as well as a lineage-specific preferential location. The proximal location of rDNA is favoured in species with multiple loci and in the two families of Hymenoptera that show the highest range of chromosome numbers: Formicidae and Vespidae. We propose that chromosome fissions have played a crucial role in the distribution pattern of rDNA loci through the evolutionary diversification of Hymenoptera. Moreover, our genomic analysis of two species, one with a single locus of rDNA and one with multiple loci, supported that loci multiplication is followed by sequence divergence. Our results provide detailed information about the number and chromosomal position of rDNA in Hymenoptera and, therefore, broaden our knowledge regarding rDNA evolutionary dynamics in insects.

Out of patterns, the euchromatic B chromosome of the grasshopper Abracris flavolineata is not enriched in high-copy repeats.

In addition to the normal set of standard (A) chromosomes, some eukaryote species harbor supernumerary (B) chromosomes. In most cases, B chromosomes show differential condensation with respect to A chromosomes and display dark C-bands of heterochromatin, and some of them are highly enriched in repetitive DNA. Here we perform a comprehensive NGS (next-generation sequencing) analysis of the repeatome in the grasshopper Abracris flavolineata aimed at uncovering the molecular composition and origin of its B chromosome. Our results have revealed that this B chromosome shows a DNA repeat content highly similar to the DNA repeat content observed for euchromatic (non-C-banded) regions of A chromosomes. Moreover, this B chromosome shows little enrichment for high-copy repeats, with only a few elements showing overabundance in B-carrying individuals compared to the 0B individuals. Consequently, the few satellite DNAs (satDNAs) mapping on the B chromosome were mostly restricted to its centromeric and telomeric regions, and they displayed much smaller bands than those observed on the A chromosomes. Our data support the intraspecific origin of the B chromosome from the longest autosome by misdivision, isochromosome formation, and additional restructuring, with accumulation of specific repeats in one or both B chromosome arms, yielding a submetacentric B. Finally, the absence of B-specific satDNAs, which are frequent in other species, along with its euchromatic nature, suggest that this B chromosome arose recently and might still be starting a heterochromatinization process. On this basis, it could be a good model to investigate the initial steps of B chromosome evolution.

B chromosomes of multiple species have intense evolutionary dynamics and accumulated genes related to important biological processes.

BACKGROUND: One of the biggest challenges in chromosome biology is to understand the occurrence and complex genetics of the extra, non-essential karyotype elements, commonly known as supernumerary or B chromosomes (Bs). The non-Mendelian inheritance and non-pairing abilities of B chromosomes make them an interesting model for genomics studies, thus bringing to bear different questions about their genetic composition, evolutionary survival, maintenance and functional role inside the cell. This study uncovers these phenomena in multiple species that we considered as representative organisms of both vertebrate and invertebrate models for B chromosome analysis. RESULTS: We sequenced the genomes of three animal species including two fishes Astyanax mexicanus and Astyanax correntinus, and a grasshopper Abracris flavolineata, each with and without Bs, and identified their B-localized genes and repeat contents. We detected unique sequences occurring exclusively on Bs and discovered various evolutionary patterns of genomic rearrangements associated to Bs. In situ hybridization and quantitative polymerase chain reactions further validated our genomic approach confirming detection of sequences on Bs. The functional annotation of B sequences showed that the B chromosome comprises regions of gene fragments, novel genes, and intact genes, which encode a diverse set of functions related to important biological processes such as metabolism, morphogenesis, reproduction, transposition, recombination, cell cycle and chromosomes functions which might be important for their evolutionary success. CONCLUSIONS: This study reveals the genomic structure, composition and function of Bs, which provide new insights for theories of B chromosome evolution. The selfish behavior of Bs seems to be favored by gained genes/sequences.

How dynamic could be the 45S rDNA cistron? An intriguing variability in a grasshopper species revealed by integration of chromosomal and genomic data.

To better understand the structure and variability of the 45S rDNA cistron and its evolutionary dynamics in grasshoppers, we performed a detailed analysis combining classical and molecular cytogenetic data with whole-genome sequencing in Abracris flavolienata, which shows extraordinary variability in the chromosomal distribution for this element. We found astonishing variability in the number and size of rDNA clusters at intra- and inter-population levels. Interestingly, FISH using distinct parts of 45S rDNA cistron (18S rDNA, 28S rDNA, and ITS1) as probes revealed a distinct number of clusters, suggesting independent mobility and amplification of the 45S rDNA components. This hypothesis is consistent with the higher genomic coverage of almost the entire cistron of 45S rDNA observed in A. flavolineata compared to other grasshoppers, besides coverage variability along the 45S rDNA cistron in the species. In addition, these differences in coverage for distinct components of the 45S rDNA cistron indicate emergence of pseudogenes evidenced by existence of truncated sequences, demonstrating the rDNA dynamics in the species. Although the chromosomal distribution of 18S rDNA was highly variable, the chromosomes 1, 3, 6, and 9 harbored rDNA clusters in all individuals with the occurrence of NOR activity in pair 9, suggesting ancestry or selective pressures to prevent pseudogenization of rDNA sequences in this chromosome pair. Additionally, small NORs and cryptic rDNA loci were observed. Finally, there was no evidence of enrichment and association of transposable elements, at least, inside or nearby rDNA cistron. These findings broaden our knowledge of rDNA dynamics, revealing an independent movement and amplification of segments of 45S rDNA cistron, which in A. flavolineata could be attributed to ectopic recombination.

High dynamism for neo-sex chromosomes: satellite DNAs reveal complex evolution in a grasshopper.

A common characteristic of sex chromosomes is the accumulation of repetitive DNA, which accounts for their diversification and degeneration. In grasshoppers, the X0 sex-determining system in males is considered ancestral. However, in some species, derived variants like neo-XY in males evolved several times independently by Robertsonian translocation. This is the case of Ronderosia bergii, in which further large pericentromeric inversion in the neo-Y also took place, making this species particularly interesting for investigating sex chromosome evolution. Here, we characterized the satellite DNAs (satDNAs) and transposable elements (TEs) of the species to investigate the quantitative differences in repeat composition between male and female genomes putatively associated with sex chromosomes. We found a total of 53 satDNA families and 56 families of TEs. The satDNAs were 13.5% more abundant in males than in females, while TEs were just 1.02% more abundant in females. These results imply differential amplification of satDNAs on neo-Y chromosome and a minor role of TEs in sex chromosome differentiation. We showed highly differentiated neo-XY sex chromosomes owing to major amplification of satDNAs in neo-Y. Furthermore, chromosomal mapping of satDNAs suggests high turnover of neo-sex chromosomes in R. bergii at the intrapopulation level, caused by multiple paracentric inversions, amplifications, and transpositions. Finally, the species is an example of the action of repetitive DNAs in the generation of variability for sex chromosomes after the suppression of recombination, and helps understand sex chromosome evolution at the intrapopulation level.

Spatial Distribution of Heterochromatin Bodies in the Nuclei of Triatoma infestans (Klug).

Constitutive heterochromatin typically exhibits low gene density and is commonly found adjacent or close to the nuclear periphery, in contrast to transcriptionally active genes concentrated in the innermost nuclear region. In Triatoma infestans cells, conspicuous constitutive heterochromatin forms deeply stained structures named chromocenters. However, to the best of our knowledge, no information exists regarding whether these chromocenters acquire a precise topology in the cell nuclei or whether their 18S rDNA, which is important for ribosome function, faces the nuclear center preferentially. In this work, the spatial distribution of fluorescent Feulgen-stained chromocenters and the distribution of their 18S rDNA was analyzed in Malpighian tubule cells of T. infestans using confocal microscopy. The chromocenters were shown to be spatially positioned relatively close to the nuclear periphery, though not adjacent to it. The variable distance between the chromocenters and the nuclear periphery suggests mobility of these bodies within the cell nuclei. The distribution of 18S rDNA at the edge of the chromocenters was not found to face the nuclear interior exclusively. Because the genome regions containing 18S rDNA in the chromocenters also face the nuclear periphery, the proximity of the chromocenters to this nuclear region is not assumed to be associated with overall gene silencing.

Uncovering the evolutionary history of neo-XY sex chromosomes in the grasshopper Ronderosia bergii (Orthoptera, Melanoplinae) through satellite DNA analysis.

BACKGROUND: Neo-sex chromosome systems arose independently multiple times in evolution, presenting the remarkable characteristic of repetitive DNAs accumulation. Among grasshoppers, occurrence of neo-XY was repeatedly noticed in Melanoplinae. Here we analyzed the most abundant tandem repeats of R. bergii (2n = 22, neo-XY♂) using deep Illumina sequencing and graph-based clustering in order to address the neo-sex chromosomes evolution. RESULTS: The analyses revealed ten families of satDNAs comprising about ~1% of the male genome, which occupied mainly C-positive regions of autosomes. Regarding the sex chromosomes, satDNAs were recorded within centromeric or interstitial regions of the neo-X chromosome and four satDNAs occurred in the neo-Y, two of them being exclusive (Rber248 and Rber299). Using a combination of probes we uncovered five well-defined cytological variants for neo-Y, originated by multiple paracentric inversions and satDNA amplification, besides fragmented neo-Y. These neo-Y variants were distinct in frequency between embryos and adult males. CONCLUSIONS: The genomic data together with cytogenetic mapping enabled us to better understand the neo-sex chromosome dynamics in grasshoppers, reinforcing differentiation of neo-X and neo-Y and revealing the occurrence of multiple additional rearrangements involved in the neo-Y evolution of R. bergii. We discussed the possible causes that led to differences in frequency for the neo-Y variants between embryos and adults. Finally we hypothesize about the role of DNA satellites in R. bergii as well as putative historical events involved in the evolution of the R. bergii neo-XY.

The repetitive DNA element BncDNA, enriched in the B chromosome of the cichlid fish Astatotilapia latifasciata, transcribes a potentially noncoding RNA.

Supernumerary chromosomes have been studied in many species of eukaryotes, including the cichlid fish, Astatotilapia latifasciata. However, there are many unanswered questions about the maintenance, inheritance, and functional aspects of supernumerary chromosomes. The cichlid family has been highlighted as a model for evolutionary studies, including those that focus on mechanisms of chromosome evolution. Individuals of A. latifasciata are known to carry up to two B heterochromatic isochromosomes that are enriched in repetitive DNA and contain few intact gene sequences. We isolated and characterized a transcriptionally active repeated DNA, called B chromosome noncoding DNA (BncDNA), highly represented across all B chromosomes of A. latifasciata. BncDNA transcripts are differentially processed among six different tissues, including the production of smaller transcripts, indicating transcriptional variation may be linked to B chromosome presence and sexual phenotype. The transcript lengths and lack of similarity with known protein/gene sequences indicate BncRNA might represent a novel long noncoding RNA family (lncRNA). The potential for interaction between BncRNA and known miRNAs were computationally predicted, resulting in the identification of possible binding of this sequence in upregulated miRNAs related to the presence of B chromosomes. In conclusion, Bnc is a transcriptionally active repetitive DNA enriched in B chromosomes with potential action over B chromosome maintenance in somatic cells and meiotic drive in gametic cells.

U1 snDNA chromosomal mapping in ten spittlebug species (Cercopidade, Auchenorrhyncha, Hemiptera).

Spittlebugs, which belong to the family Cercopidae (Auchenorrhyncha, Hemiptera), form a large group of xylem-feeding insects that are best known for causing damage to plantations and pasture grasses. The holocentric chromosomes of these insects remain poorly studied in regards to the organization of different classes of repetitive DNA. To improve chromosomal maps based on repetitive DNAs and to better understand the chromosomal organization and evolutionary dynamics of multigene families in spittlebugs, we physically mapped the U1 snRNA gene with fluorescence in situ hybridization (FISH) in 10 species of Cercopidae belonging to three different genera. All the U1 snDNA clusters were autosomal and located in interstitial position. In seven species, they were restricted to one autosome per haploid genome, while three species of the genus Mahanarva showed two clusters in two different autosomes. Although it was not possible to precisely define the ancestral location of this gene, it was possible to observe the presence of at least one cluster located in a small bivalent in all karyotypes. The karyotype stability observed in Cercopidae is also observed in respect to the distribution of U1 snDNA. Our data are discussed in light of possible mechanisms for U1 snDNA conservation and compared with the available data from other species.

The U2 snDNA Is a Useful Marker for B Chromosome Detection and Frequency Estimation in the Grasshopper Abracris flavolineata.

In this study, we describe a strategy to determine the presence of B chromosomes in the living grasshopper Abracris flavolineata by FISH using U2 snDNA as a probe in interphase hemolymph nuclei. In individuals without B chromosomes, (0B) 2 dot signals were noticed, corresponding to A complement U2 snDNA clusters. In +1B and +2B individuals, 4 or 8 additional signals were noticed, respectively. In all cases, the absence or presence of 1 or 2 B chromosomes correlated in hemolymph and in somatic or germline tissues, validating the efficiency of the marker. Our data suggest that the B chromosome of A. flavolineata is present in all somatic tissues. B-carrying individuals showed the same number of B chromosomes in germ and somatic cells, suggesting that the B is mitotically stable. The marker was used to compare B chromosome frequency in the analyzed population with a sample collected previously, in order to test for B frequency changes and differences of B chromosome prevalence among sexes, but no statistically significant differences were noticed. The identification of living animals harboring B chromosomes will be very useful in future studies of B chromosome transmission, as well as in functional studies involving RNA analysis, thus contributing to the understanding of evolutionary history and the possible role of the B chromosome in A. flavolineata.

B Chromosome Variants of the Grasshopper Xyleus discoideus angulatus Are Potentially Derived from Pericentromeric DNA.

B chromosomes, extra elements present in the karyotypes of some eukaryote species, have been described in the grasshopper Xyleus discoideus angulatus. Although some studies have proposed an autosomal origin of the B chromosome in X. d. angulatus, little is known about its repetitive DNA composition and evolutionary dynamics. The aim of the present work was to shed light on the B chromosome evolution in X. d. angulatus by cytogenetic analysis of 27 populations from Pernambuco and Ceará states (Brazil). The frequency of B chromosomes in the different populations was determined, and chromosome measurements and fluorescence in situ hybridization (FISH) with C0t-DNA and telomeric and B chromosome sequences were performed in cells from B-carrying individuals. The results revealed variations in B chromosome prevalence among the populations and showed that some B chromosomes were smaller in certain populations. FISH produced similar patterns for the C0t-DNA probe in all hybridized individuals, whereas telomeric and B chromosome probes, obtained by microdissection, exhibited variations in their distribution. These results indicate the presence of 3 morphotypes of B chromosomes in X. d. angulatus, with variation in repetitive DNA composition during their evolution. In this species, B chromosomes have an intraspecific origin and probably arose from the pericentromeric region of A chromosomes.

Are the TTAGG and TTAGGG telomeric repeats phylogenetically conserved in aculeate Hymenoptera?

Despite the (TTAGG)n telomeric repeat supposed being the ancestral DNA motif of telomeres in insects, it was repeatedly lost within some insect orders. Notably, parasitoid hymenopterans and the social wasp Metapolybia decorata (Gribodo) lack the (TTAGG)n sequence, but in other representatives of Hymenoptera, this motif was noticed, such as different ant species and the honeybee. These findings raise the question of whether the insect telomeric repeat is or not phylogenetically predominant in Hymenoptera. Thus, we evaluated the occurrence of both the (TTAGG)n sequence and the vertebrate telomere sequence (TTAGGG)n using dot-blotting hybridization in 25 aculeate species of Hymenoptera. Our results revealed the absence of (TTAGG)n sequence in all tested species, elevating the number of hymenopteran families lacking this telomeric sequence to 13 out of the 15 tested families so far. The (TTAGGG)n was not observed in any tested species. Based on our data and compiled information, we suggest that the (TTAGG)n sequence was putatively lost in the ancestor of Apocrita with at least two subsequent independent regains (in Formicidae and Apidae).

Karyotypes and Repetitive DNA Evolution in Six Species of the Genus Mahanarva (Auchenorrhyncha: Cercopidae).

Insects of the Cercopidae family are widely distributed and comprise 59 genera and 431 species in the New World. They are xylemophagous, causing losses in agricultural and pasture grasses, and are considered as emerging pests. Chromosomally, these insects have been studied by standard techniques, revealing variable diploid numbers and primarily X0 sex chromosome systems (males). We performed chromosome studies in 6 Mahanarva (Cercopidae) species using standard and differential chromosome staining as well as mapping of repetitive DNAs. Moreover, the relationship between the repetitive DNAs was analyzed at the interspecific level. A diploid chromosome number of 2n = 19,X0 was documented, with chromosomes gradually decreasing in size. Neutral or GC-rich regions were detected which varied depending on the species. Fluorescence in situ hybridization with a (TTAGG)n telomeric motif probe revealed terminal signals, matching those of the Cot DNAs obtained from each species, that were also restricted to the terminal regions of all chromosomes. Dot blot analysis with the Cot fraction from M. quadripunctata showed that at least part of the repetitive genome is shared among the 6 species. Our data highlight the conservation of chromosomal features and organization of repetitive DNAs in the genus Mahanarva, suggesting a low differentiation for chromosomes and repetitive DNAs in most of the 6 species studied.

Origin and evolution of B chromosomes in the cichlid fish Astatotilapia latifasciata based on integrated genomic analyses.

Approximately 15% of eukaryotes contain supernumerary B chromosomes. When present, B chromosomes frequently represent as much as 5% of the genome. Despite thousands of reports describing the distribution of supernumeraries in various taxa, a comprehensive theory for the origin, maintenance, and evolution of B chromosomes has not emerged. Here, we sequence the complete genomes of individual cichlid fish (Astatotilapia latifasciata) with and without B chromosomes, as well as microdissected B chromosomes, to identify DNA sequences on the B. B sequences were further analyzed through quantitative polymerase chain reaction and in situ hybridization. We find that the B chromosome contains thousands of sequences duplicated from essentially every chromosome in the ancestral karyotype. Although most genes on the B chromosome are fragmented, a few are largely intact, and we detect evidence that at least three of them are transcriptionally active. We propose a model in which the B chromosome originated early in the evolutionary history of Lake Victoria cichlids from a small fragment of one autosome. DNA sequences originating from several autosomes, including protein-coding genes and transposable elements, subsequently inserted into this proto-B. We propose that intact B chromosome genes involved with microtubule organization, kinetochore structure, recombination and progression through the cell cycle may play a role in driving the transmission of the B chromosome. Furthermore, our work suggests that karyotyping is an essential step prior to genome sequencing to avoid problems in genome assembly and analytical biases created by the presence of high copy number sequences on the B chromosome.

Repetitive DNA chromosomal organization in the cricket Cycloptiloides americanus: a case of the unusual X1X 20 sex chromosome system in Orthoptera.

A common placement for most sex chromosomes that is involved in their evolutionary histories is the accumulation of distinct classes of repetitive DNAs. Here, with the aim of understanding the poorly studied repetitive DNA organization in crickets and its possible role in sex chromosome differentiation, we characterized the chromosomes of the cricket species Cycloptiloides americanus, a species with the remarkable presence of the unusual sex chromosome system X1X20♂/X1X1X2X2♀. For these proposes, we used C-banding and mapping through the fluorescence in situ hybridization of some repetitive DNAs. The C-banding and distribution of highly and moderately repetitive DNAs (C 0t-1 DNA) varied depending of the chromosome. The greater accumulation of repetitive DNAs in the X2 chromosome was evidenced. The microsatellites were spread along entire chromosomes, but (AG)10 and (TAA)10 were less enriched, mainly in the centromeric areas. Among the multigene families, the 18S rDNA was spread throughout almost all of the chromosomes, except for pair 5 and X2, while the U2 snDNA was placed exclusively in the largest chromosome. Finally, the 5S rDNA was exclusively located in the short arms of the sex chromosomes. The obtained data reinforce the importance of chromosomal dissociation and inversion as a primary evolutionary mechanism to generate neo-sex chromosomes in the species studied, followed by the repetitive DNAs accumulation. Moreover the exclusive placement of 5S rDNA in the sex chromosomes suggests the involvement of this sequence in sex chromosome recognition throughout meiosis and, consequently, their maintenance, in addition to their avoiding degeneration.