High dose radiation exposures are rare. However, medical management of such incidents is crucial due to mortality and tissue injury risks. Rapid radiation biodosimetry of high dose accidental exposures is highly challenging, considering that they usually involve non uniform fields leading to partial body exposures. The gold standard, dicentric assay and other conventional methods have limited application in such scenarios. As an alternative, we propose Premature Chromosome Condensation combined with Fluorescent In-situ Hybridization (G0-PCC-FISH) as a promising tool for partial body exposure biodosimetry. In the present study, partial body exposures were simulated ex-vivo by mixing of uniformly exposed blood with unexposed blood in varying proportions. After G0-PCC-FISH, Dolphin\'s approach with background correction was used to provide partial body exposure dose estimates and these were compared with those obtained from conventional dicentric assay and G0-PCC-Fragment assay (conventional G0-PCC). Dispersion analysis of aberrations from partial body exposures was carried out and compared with that of whole-body exposures. The latter was inferred from a multi-donor, wide dose range calibration curve, a-priori established for whole-body exposures. With the dispersion analysis, novel multi-parametric methodology for discerning the partial body exposure from whole body exposure and accurate dose estimation has been formulated and elucidated with the help of an example. Dose and proportion dependent reduction in sensitivity and dose estimation accuracy was observed for Dicentric assay, but not in the two PCC methods. G0-PCC-FISH was found to be most accurate for the dose estimation. G0-PCC-FISH has potential to overcome the shortcomings of current available methods and can provide rapid, accurate dose estimation of partial body and high dose accidental exposures. Biological dose estimation can be useful to predict progression of disease manifestation and can help in pre-planning of appropriate & timely medical intervention.

Some types of chromosome aberrations are not easily identified by the traditional Giemsa staining. It usually needs some banding technique and skilled person\'s eye. Whole chromosome painting FISH probe can stain designated entire chromosomes or domains in metaphase chromosomes or interphase nuclei, respectively. It allows to visually identify translocations, deletions, or amplifications of specific chromosomes. Once chromosomes are stained, even non-skilled researchers can easily identify those chromosome aberrations. Whole chromosome painting FISH has higher sensitivity to detect chromosome aberrations. This chapter introduces the method for whole chromosome painting FISH staining.

The genus Gymnotus is a large monophyletic group of freshwater weakly-electric fishes, with wide distribution in Central and South America. It has 46 valid species divided into six subgenera (Gymnotus, Tijax, Tigre, Lamontianus, Tigrinus and Pantherus) with large chromosome plasticity and diploid numbers (2n) ranging from 34 to 54. Within this rich diversity, there is controversy about whether Gymnotus (Gymnotus) carapo species is a single widespread species or a complex of cryptic species. Cytogenetic studies show different diploid numbers for G. carapo species, ranging from 40 to 54 chromosomes with varied karyotypes found even between populations sharing the same 2n. Whole chromosome painting has been used in studies on fish species and recently has been used for tracking the chromosomal evolution of Gymnotus and assisting in its cytotaxonomy. Comparative genomic mapping using chromosome painting has shown more complex rearrangements in Gymnotus carapo than shown in previous studies by classical cytogenetics. These studies demonstrate that multiple chromosome pairs are involved in its chromosomal reorganization, suggesting the presence of a complex of cryptic species due to a post zygotic barrier. In the present study, metaphase chromosomes of G. carapo occidentalis \"catalão\" (GCC, 2n = 40, 30m/sm+10st/a) from the Catalão Lake, Amazonas, Brazil, were hybridized with whole chromosome probes derived from the chromosomes of G. carapo (GCA, 2n = 42, 30m/sm+12st/a). The results reveal chromosome rearrangements and a high number of repetitive DNA sites. Of the 12 pairs of G. carapo chromosomes that could be individually identified (GCA 1-3, 6, 7, 9, 14, 16 and 18-21), 8 pairs (GCA 1, 2, 6, 7, 9, 14, 20, 21) had homeology conserved in GCC. Of the GCA pairs that are grouped (GCA [4, 8], [5, 17], [10, 11] and [12, 13, 15]), most kept the number of signals in GCC (GCA [5, 17], [10, 11] and [12, 13, 15]). The remaining chromosomes are rearranged in the GCC karyotype. Analysis of both populations of the G. carapo cytotypes shows extensive karyotype reorganization. Along with previous studies, this suggests that the different cytotypes analyzed here may represent different species and supports the hypothesis that G. carapo is not a single widespread species, but a group of cryptic species.

The Neotropical genus Harttia comprises species with extensive chromosomal remodeling and distinct sex chromosome systems (SCSs). So far, three different SCSs with male heterogamety have been characterized in the group. In some species, the presence of the XX/XY1Y2 SCS is associated with a decrease in diploid numbers and several chromosomal rearrangements, although a direct relation to sex chromosome differentiation has not been shown yet. Here, we aimed to investigate the differentiation processes that have led to the establishment of the rare XX/XY1Y2 SCS and track its evolutionary history among other Harttia species. For that, four whole chromosome painting probes derived from chromosome 1 of H. torrenticola (HTO-1), chromosomes 9 and X of H. carvalhoi (HCA-9 and HCA-X), and chromosome X from H. intermontana (HIN-X) were applied in nine Harttia species. Homeologous chromosome blocks were located in Harttia species and demonstrated that Robertsonian (Rb) fusions originated HTO-1, HCA-9, and HCA-X chromosomes, while Rb fissions explain Y1 and Y2 sex chromosomes. Specifically, in H. intermontana, HCA-X, HCA-9, and the NOR-bearing chromosome demonstrated that homeologous blocks were used in the HIN-X and metacentric pair 2 origins. Consequently, diploid numbers changed between the studied species. Overall, the data also reinforce the existence of unstable genomic sites promoting chromosomal differentiation and remodeling within the genus Harttia.

Oplegnathus fasciatus and O. punctatus (Teleostei: Centrarchiformes: Oplegnathidae), are commercially important rocky reef fishes, endemic to East Asia. Both species present an X1X2Y sex chromosome system. Here, we investigated the evolutionary forces behind the origin and differentiation of these sex chromosomes, with the aim to elucidate whether they had a single or convergent origin. To achieve this, conventional and molecular cytogenetic protocols, involving the mapping of repetitive DNA markers, comparative genomic hybridization (CGH), and whole chromosome painting (WCP) were applied. Both species presented similar 2n, karyotype structure and hybridization patterns of repetitive DNA classes. 5S rDNA loci, besides being placed on the autosomal pair 22, resided in the terminal region of the long arms of both X1 chromosomes in females, and on the X1 and Y chromosomes in males. Furthermore, WCP experiments with a probe derived from the Y chromosome of O. fasciatus (OFAS-Y) entirely painted the X1 and X2 chromosomes in females and the X1, X2, and Y chromosomes in males of both species. CGH failed to reveal any sign of sequence differentiation on the Y chromosome in both species, thereby suggesting the shared early stage of neo-Y chromosome differentiation. Altogether, the present findings confirmed the origin of the X1X2Y sex chromosomes via Y-autosome centric fusion and strongly suggested their common origin.

Ionizing radiation can produce many chromosome aberrations; however, a number of aberrations are not readily detectable with a conventional Giemsa staining and require special staining methods. Moreover, an easy method to identify nondetectable chromosomal aberrations, such as symmetrical inter-chromosomal translocations, is available: whole chromosome fluorescence in situ hybridization (FISH) staining. Asymmetrical translocations such as dicentric chromosomes are known to be lethal but cells with symmetrical translocations may survive and persist in the progeny population for years. Chromosome banding techniques can also detect inter-chromosomal translocations but understanding banding patterns requires professional training and experience. Due to its clear detection by a discernible painting color, whole chromosome painting method is superior in identifying chromosome fragmentation, loss, and amplification of chromosomes. This chapter introduces whole chromosome FISH painting method to detect chromosomal aberrations such asfragments and translocations.

We present the first cytogenetic data for Lebiasina bimaculata and L. melanoguttata with the aim of (1) investigating evolutionary events within Lebiasina and their relationships with other Lebiasinidae genera and (2) checking the evolutionary relationships between Lebiasinidae and Ctenoluciidae. Both species have a diploid number 2n = 36 with similar karyotypes and microsatellite distribution patterns but present contrasting C-positive heterochromatin and CMA3+ banding patterns. The remarkable interstitial series of C-positive heterochromatin occurring in L. melanoguttata is absent in L. bimaculata. Accordingly, L. bimaculata shows the ribosomal DNA sites as the only GC-rich (CMA3+) regions, while L. melanoguttata shows evidence of a clear intercalated CMA3+ banding pattern. In addition, the multiple 5S and 18S rDNA sites in L. melanogutatta contrast with single sites present in L. bimaculata. Comparative genomic hybridization (CGH) experiments also revealed a high level of genomic differentiation between both species. A polymorphic state of a conspicuous C-positive, CMA3+, and (CGG)n band was found only to occur in L. bimaculata females, and its possible relationship with a nascent sex chromosome system is discussed. Whole chromosome painting (WCP) and CGH experiments indicate that the Lebiasina species examined and Boulengerella maculata share similar chromosomal sequences, thus supporting the relatedness between them and the evolutionary relationships between the Lebiasinidae and Ctenoluciidae families.

BACKGROUND: Species with \'young\' or nascent sex chromosomes provide unique opportunities to understand early evolutionary mechanisms (e.g. accumulation of repetitive sequences, cessation of recombination and gene loss) that drive the evolution of sex chromosomes. Among vertebrates, fishes exhibit highly diverse and a wide spectrum of sex-determining mechanisms and sex chromosomes, ranging from cryptic to highly differentiated ones, as well as, from simple to multiple sex chromosome systems. Such variability in sex chromosome morphology and composition not only exists within closely related taxa, but often within races/populations of the same species. Inside this context, the wolf fish Hoplias malabaricus offers opportunity to investigate the evolution of morphologically variable sex chromosomes within a species complex, as homomorphic to highly differentiated sex chromosome systems occur among its different karyomorphs.
METHODS: To discover various evolutionary stages of sex chromosomes and to compare their sequence composition among the wolf fish´s karyomorphs, we applied multipronged molecular cytogenetic approaches, including C-banding, repetitive DNAs mapping, Comparative Genomic Hybridization (CGH) and Whole Chromosomal Painting (WCP). Our study was able to characterize a cryptically differentiated XX/XY sex chromosome system in the karyomorph F of this species.
CONCLUSIONS: The Y chromosome was clearly identified by an interstitial heterochromatic block on the short arms, primarily composed of microsatellite motifs and retrotransposons. Additionally, CGH also identified a male specific chromosome region in the same chromosomal location, implying that the accumulation of these repeats may have initiated the Y chromosome differentiation, as well as played a critical role towards the evolution and differentiation of sex chromosomes in various karyomorphs of this species.

The Neotropical fish, Hoplias malabaricus, is one of the most cytogenetically studied fish taxon with seven distinct karyomorphs (A-G) comprising varying degrees of sex chromosome differentiation, ranging from homomorphic to highly differentiated simple and multiple sex chromosomes. Therefore, this fish offers a unique opportunity to track evolutionary mechanisms standing behind the sex chromosome evolution and differentiation. Here, we focused on a high-resolution cytogenetic characterization of the unique XX/XY1Y2 multiple sex chromosome system found in one of its karyomorphs (G). For this, we applied a suite of conventional (Giemsa-staining, C-banding) and molecular cytogenetic approaches, including fluorescence in situ hybridization FISH (with 5S and 18S rDNAs, 10 microsatellite motifs and telomeric (TTAGGG) n sequences as probes), comparative genomic hybridization (CGH), and whole chromosome painting (WCP). In addition, we performed comparative analyses with other Erythrinidae species to discover the evolutionary origin of this unique karyomorph G-specific XY1Y2 multiple sex chromosome system. WCP experiments confirmed the homology between these multiple sex chromosomes and the nascent XX/XY sex system found in the karyomorph F, but disproved a homology with those of karyomorphs A-D and other closely related species. Besides, the putative origin of such XY1Y2 system by rearrangements of several chromosome pairs from an ancestral karyotype was also highlighted. In addition, clear identification of a male-specific region on the Y1 chromosome suggested a differential pattern of repetitive sequences accumulation. The present data suggested the origin of this unique XY1Y2 sex system, revealing evidences for the high level of plasticity of sex chromosome differentiation within the Erythrinidae.

Although fishes exhibit the greatest biodiversity among the vertebrates, a large percentage of this fauna is still underexplored on evolutionary cytogenetic questions, particularly the miniature species. The Lebiasinidae family is a particular example for such case. This study is the first one presenting differential cytogenetic methods, such as C-banding, repetitive DNAs mapping, comparative genomic hybridization (CGH), and whole chromosome painting in lebiasinid species. Pyrrhulina australis and Pyrrhulina aff. australis were deeply investigated concerning their chromosomal patterns and evolutionary relationships. These species have a very similar morphology, but they can be distinguished by a longitudinal midlateral faintly dark stripe exclusive for Pyrrhulina aff. australis. Both species presented 2n = 40 chromosomes (4st +36a), without heteromorphic sex chromosomes. However, despite their morphological and karyotype resemblance, it was evidenced that both species have already gone through a significant genomic divergence, thus corresponding to distinct evolutionary units. Furthermore, to give additional support to some proposals on evolutionary relationship among Lebiasinidae with other fish families, a chromosomal comparative approach with Erythrinus erythrinus, a representative species of the Erythrinidae family, was also performed. In addition to have similar karyotype structure, mainly composed by acrocentric chromosomes, both species share uncommon genomic similarities, such as (i) syntenic location of 5S and 18S rDNA sequences; (ii) huge dispersion of multiple 5S rDNA sites in the karyotypes; and (iii) complex association between 5S rDNA and Rex3 elements. CGH experiments, despite reinforcing some shared genomic homologies, also highlighted that both Pyrrhulina and Erythrinus have a range of nonoverlapping species-specific signals. The overall chromosomal data proved to be effective markers for the cytotaxonomy and evolutionary process among Lebiasinidae fishes.