Subtelocentric Pair Research Articles

Chromosome Mapping of Ribosomal DNA Clusters in Four Leaf-cutting Ant Species of the Genus Acromyrmex, with Description of a Triploid Individual in A. laticeps

The increase of cytogenetic data in the leaf-cutting genus Acromyrmex has shown interesting contributions to the evolutionary and taxonomic approaches, including the creation of a new genus and the description of chromosomal patterns allowing comparative studies within Acromyrmex as well as between this genus and other similar taxa. The conserved chromosome number in Acromyrmex is a notable feature, with some distinguishable chromosomes among the species. In this study, we describe the diploid karyotype of Acromyrmex laticeps in addition to an individual with all the metaphases triploids, suggesting a triploid individual. To the best of our knowledge, this is the first report of polyploidy in a fungus-growing ant. Furthermore, we mapped 18S ribosomal DNA on the chromosomes of A. laticeps and three other Acromyrmex species. Among them, three species showed the ribosomal DNA cluster pattern observed in other previously studied members of this genus, with a terminal location in the largest subtelocentric pair. Meanwhile, Acromyrmex balzani had these ribosomal DNA clusters in its entire short arm. The role of triploidy in ant evolution has also been discussed. The increase in chromosomal data of leaf-cutting ants may provide significant insights into the evolution of this peculiar ant group.

First Karyotypic Insights into Potamotrygon schroederi Fernández-Yépez, 1958: Association of Different Classes of Repetitive DNA

Introduction: Currently, there are 38 valid species of freshwater stingrays, and these belong to the subfamily Potamotrygoninae. However, cytogenetic information about this group is limited, with studies mainly using classical techniques, Giemsa, and C-banding. Methods: In this study, we used classical and molecular cytogenetic techniques – mapping of 18S and 5S rDNA and simple sequence repeats (SSRs) – in order to investigate the karyotypic composition of Potamotrygon schroederi and reveal the karyoevolutionary trends of this group. Results: The species presented 2n = 66 chromosomes with 18m + 12sm + 16st + 20a, heterochromatic blocks distributed in the centromeric regions of all the chromosomes, and terminal blocks in the q arm of pairs 2 and 3. Mapping of 18S rDNA regions revealed multiple clusters on pairs 2 and 7 and a homolog of pair 24. The 5S rDNA region was found in the pericentromeric portion of the subtelocentric pair 16. Furthermore, dinucleotide SSRs sequences were found in the centromeric and terminal regions of different chromosomal pairs, with preferential accumulation in pair 17. In addition, we identified conspicuous blocks of (GATA)n and (GACA)n sequences colocalized with the 5S rDNA (pair 16). Conclusion: In general, this study corroborates the general trend of a reduction in 2n in the species of Potamotrygoninae subfamily. Moreover, we found that the location of rDNA regions is very similar among Potamotrygon species, and the SSRs accumulation in the second subtelocentric pair (17) seems to be a common trait in this genus.

Identification of cryptic species in allopatric populations of Hypostomus tietensis (Siluriformes: Loricariidae) through cytogenetics analyses

Abstract Hypostomus is the most specious genus of Hypostominae, composed of several species with high intraspecific morphological and color pattern variation, making their identification a complex issue. One of the species with problematic identification is Hypostomus tietensis that was described from a single specimen, resulting in uncertainties about its color pattern and correct identification. To assist in this context, cytogenetic analyzes were carried out in three putative populations of H. tietensis from the Upper Paraná River basin, one of them from the type locality. The three populations showed considerable cytogenetic differences, with 2n = 72 chromosomes for the population from the type locality and 2n = 76 chromosomes for the others. Terminal NORs were detected (Ag- and 18S rDNA-FISH), being simple for the type locality population (acrocentric pair 23, long arm) and the Pirapó River (subtelocentric pair 11, short arm), and multiple for Do Campo River (subtelocentric pairs 11 and 12, short and long arm, respectively). C-banding was efficient in differentiating the type locality population from the others. Cytogenetic data revealed that populations from Pirapó and Do Campo rivers, although treated until now as Hypostomus aff. tietensis, represent a cryptic species, and those morphological analyses are necessary to differentiate and for describing this new species.

Cytogenetic studies on the social parasite Acromyrmex ameliae (Formicidae: Myrmicinae: Attini) and its hosts reveal chromosome fusion in Acromyrmex

The ant Acromyrmex ameliae is a social parasite of two leaf-cutting ant subspecies: Acromyrmex subterraneus subterraneus and A. subterraneus brunneus. Cytogenetic data are available for 14 species of Acromyrmex and all of them possess 2n = 38 chromosomes. In this study, chromosome number, heterochromatin detection, and detection of AT and GC-rich blocks of colonies of A. ameliae and its hosts were carried out. Additionally, the detection of nucleolus organizer regions and 18S rDNA clusters in chromosomes of the parasite and physical mapping of telomeres were undertaken. The same chromosome number and morphology were detected for the hosts 2n = 38 (10m + 14sm + 12st + 2a), while the females and males of the social parasite A. ameliae presented 2n = 36 (10m + 16sm + 8st + 2a) and n = 18 (5m + 8sm + 4st + 1a). In both A. ameliae and its hosts, the terminal region on the short arm of the largest subtelocentric pair is heterochromatic GC-rich, and this region corresponded to the 18S rDNA clusters in the parasite. The short arms of several chromosomes were heterochromatin-rich. The telomeric probe hybridized telomeres on all chromosomes of the parasite and was not detected in intrachromosomal regions. Through a comparative cytogenetic analysis, we hypothesize that the karyotype of A. ameliae (2n = 36) originated from a chromosomal rearrangement that reduced the number of chromosomes from 38 to 36; as available data on the genus Acromyrmex show that all other species possess 38 chromosomes, representing 45% of the 33 valid species in this genus. The mechanism of the chromosome rearrangement is discussed. Thus, the chromosome number observed in A. ameliae is a derivation from the genus. Our data show variation in the chromosomal number in Acromyrmex and suggest that analyses of the karyotypes of parasite species can yield novel insights with regards to the evolution of this genus.

Chromosomal Diversification in Two Species of Pimelodus (Siluriformes: Pimelodidae): Comparative Cytogenetic Mapping of Multigene Families.

Pimelodidae has a high number of species, but cytogenetic studies are generally restricted to classical chromosomal characterization and in situ localization of ribosomal DNA (rDNA) genes. This study was developed to compare Pimelodus microstoma and Pimelodus pohli focusing on chromosomal diversification provided by the transposition of DNA sequences containing multigene families. Both species share 56 chromosomes, with centromeric and terminal heterochromatic blocks. The silver nucleolus organizer regions (Ag-NORs)/45S rDNA was located in the chromosome pair 24 for both species. The 5S rDNA sites were evidenced in the pair 8 of P. microstoma, and in the pairs 1, 17, and 18 in P. pohli. The U1 small nuclear RNA (snRNA) was located at terminal site in the first subtelocentric pair in both species. The U2 snRNA site was syntenic to 5S rDNA in non-homeologue chromosomes between analyzed species. The histones H3 and H4 were clustered in chromosome pairs 19 and 23 in P. microstoma, and 21 and 22 in P. pohli. Our study proposes that the movement of DNA sequences carrying multigene families has been driven on the chromosomal diversification of Pimelodidae. These multigene location in the genomes can explain most of the visualized chromosomal rearrangements in Pimelodidae and it is useful to understand the chromosomal changes and their distinctive karyotype formulae.

Chromosomal Localization of 18S-28S rDNA and (TTAGGG)n Sequences in Two South African Dormice of the Genus Graphiurus (Rodentia: Gliridae)

Classical cytogenetics and mapping of 18S-28S rDNA and (TTAGGG)_n sequences by fluorescence in situ hybridization (FISH) was performed on Graphiurus platyops (GPL) and Graphiurus ocularis (GOC) metaphases with the aim to characterize the genomes. In both species, inverted DAPI karyotypes showed the same diploid number, 2n = 46, and hybridization of the (TTAGGG)_n probe revealed interstitial telomeric sequences (ITSs) at the centromeres of almost all bi-armed chromosomes. FISH with the rDNA probe localized nucleolus organizer regions (NORs), at the terminal ends of the p arms of the subtelocentric pairs 16 and 17 in both species and detected additional signals on GPL8 and GOC18, 19, and 22. The species have similar karyotypes, but their chromosome pairs 18-22 differ in morphology; these are acrocentric in G. platyops, as also confirmed by C-banding, and subtelocentric in G. ocularis. These differences in pairs 18-22 were also highlighted by hybridization of the telomeric probe (TTAGGG)_n, which showed the small p arms in G. ocularis enriched with ITSs. FISH of rDNA probes detected multiple NOR loci in G. ocularis, underlining the intense evolutionary dynamics related to these genes. Although the Graphiurus species analyzed have similar karyotypes, the results on the repetitive sequences indicate a complex pattern of genomic reorganization and evolution occurring in these phylogenetically close species.

First chromosome data on Steindachneridion doceanum (Siluriformes: Pimelodidae): a critically endangered catfish endemic of the Doce River basin, Brazil

ABSTRACT The present report represents the first cytogenetic description of Steindachneridion doceanum, great catfish which is currently at high extinction risk and it is listed as threatened on the red list of the Brazilian Ministry of the Environment, also are suggested karyotype relationships with other species of the same genus endemic from other river basins. The results revealed a diploid number of 2n = 56 and the karyotype composed of 18 metacentric, 20 submetacentric, 10 subtelocentric and 8 acrocentric chromosomes (NF = 104). The AgNORs and CMA3 signals were coincident in location occupying the short arm of an acrocentric chromosome pair (25th), in a secondary constriction. The 5S rDNA genes were localized on the short arms of one subtelocentric pair. C-banding revealed terminal blocks on the short arms on many chromosomes as well as terminal positive bands at the both ends of a submetacentric pair. C banding also revealed a large heterochromatic block in the secondary constriction (25th) region that was coincident with the AgNORs sites and CMA3+ bright bands. In spite S. doceanum represent an endemic taxon, in spite their geographic isolation their cytogenetic characteristics show similarities with other species of the genus.

Cytogenetics characterization of Crenuchus spilurus (Günther, 1863): a remarkable low diploid value within family Crenuchidae (Characiformes)

Conventional and molecular cytogenetic analyses were performed in specimens of the Neotropical Crenuchus spilurus freshwater fish species from a single location (Caete River, Brazil). All specimens presented diploid values of 2n = 38 chromosomes (12 m + 4sm + 2st + 20a), the lowest reported for family Crenuchidae up to now. A single pair of nucleolar organizing regions (NORs) was detected in the subtelocentric chromosome pair no. 9 by silver-staining and fluorescence in situ hybridization (FISH) with 18S rDNA sequence-specific probe. Two pairs of 5S rRNA gene clusters were found either interstitial or terminally located in the long arms of the acrocentric chromosome pairs nos. 10 and 13. Heterochromatic regions were clearly observed in the short arms of the NOR-bearing chromosome pair and weakly-positive to the pericentromeric regions of most acrocentric chromosomes. Additionally, no sex chromosomes were identified in the surveyed specimens. Crenuchidae have signals of several mechanisms involved in karyotype diversification within this family: differential location of heterochromatin-rich regions, multiplication, and translocation of rDNA clusters, presence/absence of sex chromosomes, macrostructural changes in morphology and number of chromosomes. This variety of karyotype patterns reveals the importance of widening cytogenetic studies to more taxa for better know the chromosomal evolution occurred in this group.

Karyosystematics of an Endemic tooth-carp, Aphanius shirini (Teleostei: cyprinodontidae) from Iran

The karyological and cytological characteristics of an endemic cyprinodont fish of Iran, Aphanius shirini have been investigated for the first time by examining metaphase chromosomes spreads obtained from gill epithelial and kidney cells. The diploid chromosome number of this species is 48. The karyotype consisted of one submetacentric and 23 subtelocentric pairs of chromosomes (2Sm + 46St). The chromosome arm number (NF) is 50. Sex chromosomes were cytologically indistinguishable in this tooth-carp. Based on the present and previous reported diploid chromosome number for other cyprinodont species, it can be suggested that the diploid chromosome number of 2n = 48 is the modal number of the cyprinodont fish.

A new chromosomal race (2n=44) of Nannospalax xanthodon from Turkey (Mammalia: Rodentia)

A new karyotype for blind mole rats was recorded in Tunceli province in Eastern Turkey. The karyotype contained 44 chromosomes, including 13 biarmed pairs, 7 acrocentric pairs, and one heteromorphic pair with a submetacentric and an acrocentric homologue in the autosomal complement (FNa=69). The X chromosome was submetacentric and the Y chromosome medium-sized subtelocentric (FN=73). Distinct dark centromeric C-bands were observed on most of the biarmed and three pairs of the acrocentric autosomes. The NORs were detected on short arms of three subtelocentric pairs and one acrocentric pair of autosomes. The diploid number of chromosomes and the karyotype characteristics observed are obviously unique among hitherto studied populations of blind mole rats and the complement can be evaluated as a new chromosome race of Nannospalax xanthodon. The distribution ranges of individual chromosome races of the species recorded in Eastern Anatolia are revised and possible interracial hybridization is discussed in respect of the finding of a new race.

G-banding of the genus Rattus (rodent, Muridae) in Tunisia, occurrence of different banding of the sexual chromosome Y in the norvegicus species

The two species of rats Rattus rattus and Rattus norvegicus are the only representatives of the genus Rattus in Tunisia. R. rattus is represented by a karyotye 2n = 38 to 39 chromosomes, it belongs to the oceanic type, whereas R. norvegicus has a karyotype of 2n = 42 chromosomes. The G-bands produced by these two species are specific ; the ideogram representation of the chromosomes shows 136 bands for R. rattus and 189 bands for R. norvegicus, the number of the bands for each species is lower than those identified in Europe. Based on the comparison of the various G-banding profiles, we can confirm that for R. rattus, the integration of the acrocentrics pairs 4/ 7 and 11/ 12 by two robertsonian fusions and the two pericentric inversions of acrocentric pairs 1 and 9, who have infected the asian type, give respectively the two large metacentric pairs and the two subtelocentric pairs (1 and 9) characteristic of the oceanic type and the B chro- mosome is completely heterochromatic. For R. norvegicus, the change of the position between the dark and clear bands of the sexual chromosome Y is the remarkable opposite of the one identified in Europe.

First Description of supernumerary Chromosomes in Ictaluruspunctatus Rafinesque 1818 Reveals Active Ribosomal Genes in the B Complement.

The North American channel catfish Ictalurus punctatus Rafinesque 1818 is cultivated in the United States, Asia and Brazilian fish farms, and also utilized as a model species in aquaculture and genetic studies. In this work, cytogenetic analysis of . punctatus from Brazilian aquaculture revealed for the first time the presence of extra elements (supernumerary or B chromosomes) in this species. These elements were characterized as dot-like micro B chromosomes and were found in three individuals (varying from 0 to 1) and in one individual with higher incidence per cell (varying from 0 to 5; mean number of Bs per cell = 2.01). More specific cytogenetic techniques in this individual revealed 58 A chromosomes (standard complement) containing heterochromatic bands in the centromeric regions, a single Ag-NOR in a subtelocentric pair (also positive for 18S rDNA using the FISH technique) and multiple 5S rDNA clusters in three different subtelocentric chromosomes. Four B chromosomes were entirely Ag-NOR positive (also fully heterochromatic) and three presented 18S rDNA clusters by FISH. The occurrence of Ag-NOR and 18S ribosomal genes in both A and B chromosome complements may indicate an intraspecific origin for these extra chromosomes. Additionally, the terminal location of 18S ribosomal clusters in the Ag-NOR-bearing chromosomes and the presence of active NOR in the B chromosomes suggested that breakage events may be related to a possible recent origin of these extra elements. We suggest this data may be useful as cytogenetic information for future elucidation of the composition, origin and evolution of extra chromosomes in fishes.

Analysis of the histone cluster in Senegalese sole (Solea senegalensis): evidence for a divergent evolution of two canonical histone clusters.

The Senegalese sole (Solea senegalensis) is commercially very important and a priority species for aquaculture product diversification. The main histone cluster was identified within two BAC clones. However, two replacement histones (H1.0 and H3.3) were found in another BAC clone. Different types of canonical histones H2A and H2B were found within the same species for the first time. Phylogenetic analysis demonstrated that the different types of H1, H2A, and H2B histones were all more similar to each other than to canonical histones from other species. The canonical histone H3 of S. senegalensis differs from subtypes H3.1 and H3.2 in humans at the site of residue 96, where a serine is found instead of an alanine. This same polymorphism has been found only in Danio rerio. The karyotype of S. senegalensis comprises 21 pairs of chromosomes, distributed in 3 metacentric pairs, 2 submetacentric pairs, 4 subtelocentric pairs, and 12 acrocentric pairs. The two BAC clones that contain the clusters of canonical histones were both mapped on the largest metacentric pair, and mFISH analysis confirmed the co-location with the dmrt1 gene in that pair. Three chromosome markers have been identified which, in addition to those previously described, account for 18 chromosome pairs in S. senegalensis.

Estructura cariotípica de dos poblaciones de <i>Rhamdia quelen</i> (Pisces, Siluriformes) de los Ríos Paraná y Paraguay (Argentina)

Se analizaron 13 ejemplares de Rhamdia quelen (4 hembras y 9 machos) capturados en el río Paraná y 7 ejemplares (3 hembras y 4 machos) del río Paraguay. En total se registraron 244 metafases. Los especimenes mostraron un número diploide de 58 cromosomas. Con una formula cariotípica de 46M/SM + 12ST, el número fundamental (FN) fue de 116. No se registraron diferencias entre ambas poblaciones. No se evidenció la ocurrencia de heteromorfismo cromosómico relacionado con los sexos. En ambas poblaciones, las regiones organizadoras de nucléolos (NOR) se localizaron en la región terminal del brazo corto de un par subtelocéntrico (par 27). Los cromosomas del complemento A mostraron bandas C+ distribuidas principalmente sobre las NOR, en las regiones terminales y pericentroméricas de unos pocos cromosomas. Se concluye que las poblaciones de R. quelen analizadas antuvieron su macro-estructura cariotípica conservada con respecto al número diploide de 58, con cromosomas de dos brazos, región organizadora de nucléolos de tipo simple y un patrón pobre de bandas C.

Karyosystematics of Kol tooth-carp, Aphanius darabensis (Teleostei: Cyprinodontidae)

The karyological and cytological characteristics of an endemic cyprinodont fish of Iran, Aphanius darabensis Esmaeili, Teimori, Gholami & Reichenbacher, 2014 have been investigated for the first time by examining metaphase chromosomes spreads obtained from gill epithelial and kidney cells. The diploid chromosome number of A. darabensis is 48. The karyotype consisted of five submetacentric and 19 subtelocentric pairs of chromosomes (5sm+19st). The fundamental number (FN) is 58. Sex chromosomes were cytologically indistinguishable in this tooth-carp. According to this study and previous karyological reports from other cyprinodont species, it can be suggested that the diploid number (2n=48) is common amongst cyprinodont fishes. These results can be used as basic informations in population studies and management and conservation programs.

Karyotypes ofCualac tesselatusandFloridichthys carpio: comments on the phylogenetic distribution of multiple sex chromosomes in North American cyprinodontids

Abstract Close relatives of the small genera Cualac (one species) and Floridichthys (two species) in the family Cyprinodontidae show sexual differences in chromosome number, with females having 2n = 48 and males having 2n = 47 with a large Y chromosome representing chromosomal fusion. However, karyotypic analysis of both sexes in F. carpio and C. tesselatus revealed the typical cyprinodontid karyotype, with 2n = 48, one normal-sized metacentric to submetacentric chromosome pair, and a graded series of 23 acrocentric or subtelocentric pairs. The phylogenetic pattern suggests that sexually dimorphic karyotypes have arisen independently on two separate occasions in a group that otherwise shows unusually high stability in chromosome morphology.

Cytogenetic data on six leafcutter ants of the genus Acromyrmex Mayr, 1865 (Hymenoptera, Formicidae, Myrmicinae): insights into chromosome evolution and taxonomic implications

Cytogenetic data for the genus Acromyrmex Mayr, 1865 are available, to date, for a few species from Brazil and Uruguay, which have uniform chromosome numbers (2n = 38). The recent cytogenetic data of Acromyrmex striatus (Roger, 1863), including its banding patterns, showed a distinct karyotype (2n = 22), similar to earlier studied Atta Fabricius, 1804 species. Karyological data are still scarce for the leafcutter ants and many gaps are still present for a proper understanding of this group. Therefore, this study aimed at increasing cytogenetic knowledge of the genus through the characterization of other six species: Acromyrmex balzani (Emery, 1890), Acromyrmex coronatus Fabricius, 1804, Acromyrmex disciger (Mayr, 1887), Acromyrmex echinatior (Forel, 1899), Acromyrmex niger (Smith, 1858) and Acromyrmex rugosus (Smith, 1858), all of which were collected in Minas Gerais – Brazil, except for Acromyrmex echinatior which was collected in Barro Colorado – Panama. The number and morphology of the chromosomes were studied and the following banding techniques were applied: C-banding, fluorochromes CMA3 and DAPI, as well as the detection of 45S rDNA using FISH technique. All the six species had the same chromosome number observed for already studied species, i.e. 2n = 38. Acromyrmex balzani had a different karyotype compared with other species mainly due to the first metacentric pair. The heterochromatin distribution also showed interspecific variation. Nevertheless, all the studied species had a pair of bands in the short arm of the first subtelocentric pair. The fluorochrome CMA3 visualized bands in the short arm of the first subtelocentric pair for all the six species, while Acromyrmex rugosus and Acromyrmex niger also demonstrated in the other chromosomes. The AT-rich regions with differential staining using DAPI were not observed. 45S ribosomal genes were identified by FISH in the short arm of the first subtelocentric pair in Acromyrmex coronatus, Acromyrmex disciger and Acromyrmex niger. The uniform chromosome number in the genus Acromyrmex (2n = 38) suggests that Acromyrmex striatus (2n = 22) should be transferred to a new genus. Other aspects of the chromosome evolution in ants are also discussed.

KARYOLOGY OF MUGIL LIZA AND M. CUREMA FROM VENEZUELA

Karyotypes of Mugil liza and M. curema from Venezuela were studied. M. liza karyotype is 2n=48 acrocentric chromosomes, but M. curema is 2n=24, containing one submetacentric and 11 metacentric pairs. The chromosomic complement described for M. liza constitutes the first report for the speciesand coincides with the modal karyotype (2n=48) in the Mugilidae family. Venezuelan M. curema karyotype differs in number and shape from M. curema from Louisiana (USA), which has been reported to have a complement 2n=28 (10 metacentric pairs, two subtelocentric pairs and two acrocentric pairs).

Diversity of Sex Chromosome Systems in Ancistrini (Loricariidae, Hypostominae): ZZ/ZW in Ancistrus taunayi Miranda Ribeiro, 1918

The karyotype of the Ancistrini catfish Ancistrus taunayi was analyzed by conventional (Giemsa staining, AgNOR staining and C-banding) and molecular cytogenetic (5S and 18S rDNA-FISH) methods. The diploid chromosome number was 2n = 50 (22 metacentrics + 10 submetacentrics + 10 subtelocentrics + 8 acrocentrics) for both sexes. A single NOR-bearing acrocentric chromosome pair (No. 24) was detected after Ag-staining and 18S rDNA-FISH, while 5S rDNA was found only in the subtelocentric pair No. 21. Conspicuous GC-rich heterochromatin blocks corresponded to the NOR sites and were also observed in the distal regions of the acrocentric chromosome pairs Nos. 22 and 25. Chromosome pair No. 22 differed between males and females; in males, only a small interstitial block of GC-rich heterochromatin was present in both chromosomes, whereas in females, 2 blocks of GC-rich heterochromatin flanked a euchromatic region in one of the homologues, suggesting the occurrence of a ZZ/ZW sex chromosome system. Two mechanisms for the origin and evolution of this simple ZZ/ZW sex chromosome system in A. taunayi are proposed: (1) a paracentric inversion followed by amplification of the proximal heterochromatin and (2) amplification of the interstitial heterochromatin followed by a paracentric inversion. Although ZZ/ZW systems have already been described for other Ancistrus species, our results do not show the same pattern, suggesting an independent origin.

Karyological analyses of Pseudhymenochirus merlini and Hymenochirus boettgeri provide new insights into the chromosome evolution in the anuran family Pipidae

Karyological studies continue to clarify the evolutionary history of many taxa. We report here on the first chromosome survey of the frog Pseudhymenochirus merlini (Pipidae) in comparison with the related species Hymenochirus boettgeri.In addition to the 2n=20 regular (A) chromosomes (eight metacentric and two subtelocentric pairs), H. boettgeri has an accessory B chromosome in about half of the scored karyotypes (2n=21). P. merlini showed a karyotype of 2n=20 chromosomes, differing from the set of A chromosomes of H. boettgeri by three inversions.Other differences are in the location of the nucleolus organizer regions (NORs), on the 10th chromosome pair in P. merlini instead of the 4th pair in H. boettgeri. Heterochromatin is less abundant in P. merlini, and localized in centromeric and telomeric regions of all chromosomes. The patterns in H. boettgeri resemble those of another pipid species, Silurana tropicalis, as to chromosome number (2n=20), morphology, and G-banding patterns. We propose that these characteristics are symplesiomorphic in Pipidae, allowing us to present a scenario for the chromosome evolution in this family. Our results identify a third mechanism of increase in chromosome numbers among pipid frog species, the addition of B chromosomes in Hymenochirus, besides the known processes of polyploidization in Xenopus/Silurana, and centric fission in Pipa.