Ontogenetic allometric coefficient changes: implications of diet shift and morphometric traits in Hoplias malabaricus (Bloch) (Characiforme, Erythrinidae)

This study evaluated the relationship between body size and digestive tract characteristics of the important predatory freshwater fish Hoplias malabaricus , which is widely distributed in South America. The allometric coefficients were calculated for the mass and standard length ( L _S) relationships for two different L _S groups: (1) between 20 and 100 mm (characterized as insectivores) and (2) >100 mm (characterized as piscivores). Differential growth measured from the allometric coefficient, b , between the insectivore ( b < 3) and the piscivore ( b > 3) groups was detected. Anterior intestine length and pyloric caeca zone length showed significant differences between groups. Two complementary hypotheses were developed to explain the differential growth: (1) H. malabaricus has a digestive tract adapted to a piscivorous diet, which is independent of its ontogenetic stage of development, and (2) the negative allometry observed in group 1 individuals agrees with a general behavioural strategy, allowing individuals to grow in L _S during a shorter period of time.     

Ultrastructure of spermatogenic cells and spermatozoa in Hoplias malabaricus (Teleostei, Characiformes, Erythrinidae)

The differentiation of spermatids in Hoplias malabaricus is characterized by chromatin compaction, flagellum development, nuclear rotation, nuclear fossa formation, and excess cytoplasm elimination. In the resulting spermatozoon, the head is round and the nucleus contains chromatin compacted in thick filaments, peripherically arranged, to a central electron-lucent area. The acrosome is absent. The nuclear fossa is eccentric but not pronounced. The proximal centriole penetrates it and is oblique to the flagellum. The long midpiece has several converging elongate vesicles, forming membranous hoops in the initial segment of the flagellum, but has no cytoplasmic channel. The mitochondria are elongate and branched or C-shaped and located around the initial segment of the axoneme. The lateral flagellum does not show lateral projections. The ultrastructural characteristics of H.malabaricus spermatozoa are similar to the Cypriniformes.     

Hepatocytes primary culture from the Neotropical fish, trahira Hoplias malabaricus (Bloch)

Hepatocytes were isolated and cultured from Hoplias malabaricus using a non-enzymatic protocol. From 6·9 to 9·4 × 10⁷ hepatocytes g⁻¹ of liver were isolated and cell viability was 65–85%. Fish hepatocytes attached well to untreated polystyrene flasks (Corning) as well as on treated surfaces with extracellular matrix proteins, i.e. fibronectin, matrigel or type I collagen coatings. Cells rapidly started migrating and reorganizing in cord-like groups after seeding, with spaces similar to bile canaliculi in vivo . Prominent nucleolus and euchromatin-rich rounded nucleus, abundant mitochondria, well-developed rough endoplasmic reticulum, polysomes and Golgi apparatus were seen under ultrastructural analysis. Cells remained functional and metabolically active after 6 days in culture. The protocol established in the current work provides the basis for further studies of native fishes for accurate in vitro toxicological studies.     

A tandemly repetitive centromeric DNA sequence of the fish Hoplias malabaricus (Characiformes: Erythrinidae) is derived from 5S rDNA

A substantial fraction of the eukaryotic genome consists of repetitive DNA sequences that include satellites, minisatellites, microsatellites, and transposable elements. Although extensively studied for the past three decades, the molecular forces that generate, propagate and maintain repetitive DNAs in the genomes are still discussed. To further understand the dynamics and the mechanisms of evolution of repetitive DNAs in vertebrate genome, we searched for repetitive sequences in the genome of the fish species Hoplias malabaricus. A satellite sequence, named 5SHindIII-DNA, which has a conspicuous similarity with 5S rRNA genes and spacers was identified. FISH experiments showed that the 5S rRNA bona fide gene repeats were clustered in the interstitial position of two chromosome pairs of H. malabaricus, while the satellite 5SHindIII-DNA sequences were clustered in the centromeric position in nine chromosome pairs of the species. The presence of the 5SHindIII-DNA sequences in the centromeres of several chromosomes indicates that this satellite family probably escaped from the selective pressure that maintains the structure and organization of the 5S rDNA repeats and become disperse into the genome. Although it is not feasible to explain how this sequence has been maintained in the centromeric regions, it is possible to hypothesize that it may be involved in some structural or functional role of the centromere organization.     

Molecular cytogenetic analysis and the establishment of a cell culture in the fish species Hollandichthys multifasciatus (Eigenmann & Norris, 1900) (Characiformes,Characidae)

Hollandichthys is a fish genus of the family Characidae that was until recently considered to be monotypic, with cytogenetic, morphological, and molecular data being restricted to a few local populations. In the present study, the karyotype of a population of Hollandichthys multifasciatus was analyzed using classical and molecular cytogenetic approaches for the investigation of potential markers that could provide new perspectives on the cytotaxonomy. H. multifasciatus presented a diploid number of 2n=50 chromosomes and a karyotype formula of 8m+10sm+32st. A single pair of chromosomes presented Ag-NORs signals, which coincided with the 18S rDNA sites visualized by FISH, whilst the 5S rDNA sequences were mapped in two chromosome pairs. The distribution of the U snRNA genes was mapped on the Hollandichthys chromosomes for the first time, with the probes revealing the presence of the U1 snDNA on the chromosomes of pair 20, U2 on pairs 6 and 19, U4 on pair 16, and U6 on the chromosomes of pair 11. The results of the present study indicated karyotypic differences in comparison with the other populations of H. multifasciatus studied previously, reinforcing the need for further research to identify isolated populations or the potential existence of cryptic Hollandichthys species.     

Cardiac function of two ecologically distinct Neotropical freshwater fish: Curimbata, Prochilodus lineatus (Teleostei, Prochilodontidae), and trahira, Hoplias malabaricus (Teleostei, Erythrinidae)

An isometric muscle preparation was used to investigate the importance of the ventricular sarcoplasmic reticulum (SR) and extracellular Ca(2+) (2.5 up to 10.5 mM) to force generation at 25 degrees C (acclimation temperature) in two ecologically distinct Neotropical teleost fish: Curimbata (active species), and trahira (sedentary species). The post-rest force was studied with and without 10 muM ryanodine in the medium. The positive inotropism observed for both species in response to increases on extracellular Ca(2+) reflected a greater Ca(2+) influx through sarcolemma, as well as an increase in Ca(2+) liberation from the SR by the Ca(2+)-induced Ca(2+) release mechanism. The significant post-rest potentiation recorded for the curimbata and trahira control preparations (3.22+/-0.24 to 6.55+/-0.77 mN mm(-2) and 0.74+/-0.07 to 2.26+/-0.26 mN mm(-2), respectively), was completely inhibited by the addition of ryanodine to the bathing medium, suggesting a potential functionality of SR for both species. Considering the differences in these species habitats, modes of life and levels of activity and the fact of a probable SR Ca(2+) cycling in a physiological temperature, we suggest that the functionality of the SR in these species is probably related to their phylogeny.     

New and previously described species of Urocleidoides (Monogenoidea: Dactylogyridae) infecting the gills and nasal cavities of Hoplias malabaricus (Characiformes: Erythrinidae) from Brazil

During an investigation of the biodiversity and structure of parasite communities among native populations of the erythrinid fish Hoplias malabaricus (Characiformes) from 7 rivers in Brazil, the following monogenoidean (Dactylogyridae) species were found: Urocleidoides malabaricusi n. sp., Urocleidoides naris n. sp., Urocleidoides cuiabai n. sp., Urocleidoides brasiliensis n. sp., and Urocleidoides eremitus Kritsky, Thatcher, and Boeger, 1986. Specimens considered as a new genus of Dactylogyridae were also found. The new species are mainly distinguished from the other 14 species of Urocleidoides in the general morphology of their copulatory and anchors/bar complexes. These new species and the new dactylogyrid genus are described, as well as supplemental observations and new illustrations of U. eremitus are provided. The present findings expand the known geographical distribution of species of Urocleidoides to southeastern and midwestern Brazil. A high speciation of this genus in the tropics is hypothesized, and it is briefly discussed.     

Cytogenetic divergence in two sympatric fish species of the genus Astyanax Baird and Girard, 1854 (Characiformes,Characidae) from northeastern Brazil

The fish genus Astyanax is widespread throughout the Neotropical region and is one of the most species-rich genera of the Characiformes. Cytogenetic studies of Astyanax have revealed marked intra- and interspecific diversity, with the identification of various species complexes. In this report, we describe the karyotypic structure of two sympatric species of Astyanax (Astyanax sp. and Astyanax aff. fasciatus) from the Middle Contas River basin in the northeastern Brazilian state of Bahia. Both species had 2n = 48 but differed in their karyotypic formulae. Small heterochromatic blocks and multiple nucleolar organizer regions (NORs) were identified in both species. Terminal CMA₃⁺/DAPI⁻ signals were observed in Astyanax sp. and A. aff. fasciatus, mostly coincident with NORs. These results show that chromosomal markers can be used to identify species in this fish complex. These markers can provide useful information for evolutionary studies and investigations on the mechanisms of chromosomal diversity in Astyanax.     

Geological subsidence, river capture, and cladogenesis of galaxiid fish lineages in central New Zealand

We used mtDNA and isozyme analysis of a freshwater fish, Galaxias divergens (Osmeriformes: Galaxiidae), to test a hypothesis of drainage evolution in South Island, New Zealand. Geological evidence indicates that the presently north-flowing Kaituna River branch of the Pelorus River system once flowed south into the Wairau River system. The subsequent flow-reversal is thought to have resulted from Pleistocene subsidence in central New Zealand. mtDNA sequence data corroborated this geological hypothesis: rivers draining into Pelorus Sound were found to retain a genetic lineage of G. divergens that is otherwise restricted to the Wairau River system and adjacent coastal drainages (based on current sampling). Other sampled drainages in northern South Island and southern North Island were found to house lineages that were highly divergent from the Wairau–Pelorus clade. Isozyme data yielded groupings based on fixed differences that were largely congruent with mtDNA clades. Standard molecular calibrations suggest that vicariant isolation of Pelorus and Wairau systems (drainage reversal) occurred in the mid-Pleistocene rather than the late Pleistocene as suggested by geology. Future multidisciplinary analyses will aim to improve our understanding of geological and molecular evolutionary rates.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society, 2006, 88 , 367–376.     

Genetic differentiation among distinct karyomorphs of the wolf fish Hoplias malabaricus species complex (Characiformes,Erythrinidae) and report of unusual hybridization with natural triploidy

In this study, genetic differentiation between karyomorphs A (2n = 42) and D (2n = 39/40) of the wolf fish Hoplias malabaricus, which is comprised of several cryptic species that present a wide variety of diploid chromosome numbers and sex chromosome systems, resulting in the identification of seven distinct karyomorphs (A–G), was investigated using a combination of molecular and cytogenetic tools. Deep sequence divergences for both karyomorphs were observed and indicate a long period of reproductive isolation between karyomorphs A and D. Additionally, one individual with 61 chromosomes was identified, which, as far as is known, is the first case of natural triploidy resulting from the hybridization between these highly differentiated karyomorphs of H. malabaricus. Molecular and cytogenetic analyses revealed that this allotriploid specimen carries two sets of maternal chromosomes from karyomorph D (2n = 40) and one set of chromosomes from karyomorph A (n = 21). Moreover, ribosomal sites and active nucleolus organizer regions from both parental contributors were found in the triploid hybrid. Considering the significant genetic distances between karyomorphs A and D, one of the primary reasons for the lack of recurrent reports of hybridization in the H. malabaricus species complex may be due to post‐zygotic barriers, such as hybrid sterility or unviability.     

Phylogenetic relationships, diversification and biogeography in Neotropical Brotogeris parakeets

Aim We present a molecular phylogenetic analysis of Brotogeris (Psittacidae) using several distinct and complementary approaches: we test the monophyly of the genus, delineate the basal taxa within it, uncover their phylogenetic relationships, and finally, based on these results, we perform temporal and spatial comparative analyses to help elucidate the historical biogeography of the Neotropical region. Location Neotropical lowlands, including dry and humid forests. Methods Phylogenetic relationships within Brotogeris were investigated using the complete sequences of the mitochondrial genes cyt b and ND2, and partial sequences of the nuclear intron 7 of the gene for Beta Fibrinogen for all eight species and 12 of the 17 taxa recognized within the genus (total of 63 individuals). In order to delinetae the basal taxa within the genus we used both molecular and plumage variation, the latter being based on the examination of 597 skin specimens. Dates of divergence and confidence intervals were estimated using penalized likelihood. Spatial and temporal comparative analyses were performed including several closely related parrot genera. Results Brotogeris was found to be a monophyletic genus, sister to Myiopsitta. The phylogenetic analyses recovered eight well‐supported clades representing the recognized biological species. Although some described subspecies are diagnosably distinct based on morphology, there was generally little intraspecific mtDNA variation. The Amazonian species had different phylogenetic affinities and did not group in a monophyletic clade. Brotogeris diversification took place during the last 6 Myr, the same time‐frame as previously found for Pionus and Pyrilia. Main conclusions The biogeographical history of Brotogeris implies a dynamic history for South American biomes since the Pliocene. It corroborates the idea that the geological evolution of Amazonia has been important in shaping its biodiversity, argues against the idea that the region has been environmentally stable during the Quaternary, and suggests dynamic interactions between wet and dry forest habitats in South America, with representatives of the Amazonian biota having several independent close relationships with taxa endemic to other biomes.     

Transcontinental dispersal,ecological opportunity and origins of an adaptive radiation in the Neotropical catfish genus Hypostomus (Siluriformes: Loricariidae)

Ecological opportunity is often proposed as a driver of accelerated diversification, but evidence has been largely derived from either contemporary island radiations or the fossil record. Here, we investigate the potential influence of ecological opportunity on a transcontinental radiation of South American freshwater fishes. We generate a species‐dense, time‐calibrated molecular phylogeny for the suckermouth armored catfish subfamily Hypostominae, with a focus on the species‐rich and geographically widespread genus Hypostomus. We use the resulting chronogram to estimate ancestral geographical ranges, infer historical rates of cladogenesis and diversification in habitat and body size and shape, and test the hypothesis that invasions of previously unoccupied river drainages accelerated evolution and contributed to adaptive radiation. Both the subfamily Hypostominae and the included genus Hypostomus originated in the Amazon/Orinoco ecoregion. Hypostomus subsequently dispersed throughout tropical South America east of the Andes Mountains. Consequent to invasion of the peripheral, low‐diversity Paraná River basin in southeastern Brazil approximately 12.5 Mya, Paraná lineages of Hypostomus, experienced increased rates of cladogenesis and ecological and morphological diversification. Contemporary lineages of Paraná Hypostomus are less species rich but more phenotypically diverse than their congeners elsewhere. Accelerated speciation and morphological diversification rates within Paraná basin Hypostomus are consistent with adaptive radiation. The geographical remoteness of the Paraná River basin, its recent history of marine incursion, and its continuing exclusion of many species that are widespread in other tropical South American rivers suggest that ecological opportunity played an important role in facilitating the observed accelerations in diversification.     

Molecular genetics of Rhabdomys pumilio subspecies boundaries: mtDNA phylogeography and karyotypic analysis by fluorescence in situ hybridization

The phylogeography of the African four-striped mouse, Rhabdomys pumilio, was investigated using complete sequences of the mtDNA cytochrome b gene (1140 bp) and a combination of fluorescence in situ hybridization (FISH) and conventional cytogenetic banding techniques (G- and C-banding). Two cytotypes (2n=46 and 2n=48) were identified by cytogenetic analysis. There is no evidence of diploid number variation within populations, difference in gross chromosome morphology or of subtle interchromosomal rearrangements at levels detected by ZOO-FISH. Analysis of the mtDNA cytochrome b resulted in two major lineages that correspond roughly to the xeric and mesic biotic zones of southern Africa. One mtDNA clade comprises specimens with 2n=48 and the other representatives of two cytotypes (2n=48 and 2n=46). The mean sequence divergence (12%, range 8.3–15.6%) separating the two mtDNA clades is comparable to among-species variation within murid genera suggesting their recognition as distinct species, the prior names for which would be R. dilectus and R. pumilio. Low sequence divergences and the diploid number dichotomy within the mesic lineage support the recognition of two subspecies corresponding to R. d. dilectus (2n=46) and R. d. chakae (2n=48). Our data do not support subspecific delimitation within the nominate, R. pumilio. Molecular dating places cladogenesis of the two putative species at less than five million years, a period characterised by extensive climatic oscillations which are thought to have resulted in habitat fragmentation throughout much of the species range.     

Phylogenetic systematics and historical biogeography of the Neotropical electric fish Sternopygus (Teleostei: Gymnotiformes)

Interrelationships among 10 extant species of the Neotropical electric fish Sternopygus are inferred from phylogenetic analysis of 66 morphological characters, including features of pigmentation, body proportions, meristics and osteology. A total of 287 lots containing 677 specimens were examined. The important findings of this study are: (1) S. branco is the most basal species unique among congeners in being restricted to whitewater rivers in the Central Amazon Basin, (2) S. sp. ‘cau’ from the Rio Caura of Venezuela is the sister taxon to (S. obtusirostris + S. astrabes), (3) S. castroi is a junior synonym of S. astrabes, (4) S. macrurus is the sister taxon to (S. arenatus + S. xingu + S. aequilabiatus species group) and (5) S. arenatus is the sister taxon to (S. xingu + S. aequilabiatus species group). A key to the adults of Sternopygus species is provided. Several features of S. astrabes previously thought to be plesiomorphic are now considered derived, including: short body cavity, paedomorphic cranial osteology, and the habitat restriction to terra firme streams. Sternopygus species assemblages in the Pacific (trans‐Andean) and Atlantic (cis‐Andean) slopes of northwestern South America are not monophyletic and do not result exclusively from local or regional radiations. The clade composed of S. macrurus, S. arenatus, S. xingu and the S. aequilabiatus species group is inferred to predate the Middle Miocene uplift of the Eastern Cordillera (c. 11.8–12.2 Ma). As currently recognized S. macrurus is the most widely distributed and most eurytopic gymnotiform species, inhabiting all hydrogeographical regions of tropical South America and most lowland aquatic habitats. Other Sternopygus species have much more restricted geographic and ecological distributions. Perceptions of phylogenetic patterns in Sternopygus are shown to be highly sensitive to taxon sampling.     

The vanishing clone: karyotypic evidence for extensive intraclonal genetic variation in the peach potato aphid,Myzus persicae (Hemiptera: Aphididae)

Analysis of holocentric mitotic metaphase chromosomes of the peach‐potato aphid Myzus persicae (Sulzer) clone 33H revealed different chromosome numbers, ranging from 12 to 17 within each embryo, in contrast to the standard karyotype of this species (2n = 12). Chromosome length measurements revealed that the observed chromosomal mosaicism is the result of recurrent fragmentations of chromosomes X, 1 and 3 because of fragile sites or hot spots of recombination. Fluorescent in situ hybridization experiments showed that X chromosomes were frequently involved in recurrent fragmentations, in particular their telomeric end opposite to the nucleolar organizer region. Experiments to induce males showed that M. persicae clone 33H is obligately parthenogenetic. The reproduction by apomictic parthenogenesis, together with a high telomerase expression that stabilized the chromosomes involved in the fragmentations observed in the M. persicae clone 33H, appears to favour the stabilization of the observed chromosome instability. © 2011 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 105 , 350–358.     

Molecular phylogeny and timing of radiation in Lessonia (Phaeophyceae,Laminariales)

The genus Lessonia Bory de Saint‐Vincent is distributed solely in the Southern Hemisphere, with four species found in South America and five in Australasia. Our goal was to determine the evolutionary relationships between the Lessonia species of the two disjunct areas and hypothesize dispersal patterns and timing of radiation. We combined mitochondria, plastid and nuclear markers in a comprehensive dataset from multiple individuals per known species. Furthermore, for some species we added samples from multiple populations to take account of their widespread distribution over known bioregions. In all analyses the Australasian Lessonia formed a monophyletic group and in most analyses the South American species form a paraphyletic grade. Delimitations of the accepted species are highly supported except for L. variegata J. Agardh and L. nigrescens Bory de Saint‐Vincent. We showed for the first time four lineages for the New Zealand endemic Lessonia variegata with an unexpected high level of genetic differentiation. Our analysis cannot resolve whether the dispersal of the common ancestor of this genus was from the Americas to Australasia or vice versa. Molecular clock analysis suggested that a sudden radiation took place in Australasia c. 3.5 Mya when almost all Australasian species diverged within a time frame of only 35 000 years.