Fine structures of oocyte envelopes of three related cobitid species in the genus Iksookimia (Cobitidae)

Fine structures of the oocyte envelope (zona radiata) in three spined loaches, Iksookimia longicorpus, I. hugowolfeldi, and I. yongdokensis, are examined and compared. The zona radiata surrounding the oocyte is composed of two layers, a zona radiata externa, which comprised the outermost zona radiata (Z₁) of a homogeneous less electron dense single layer and the middle zona radiata (Z₂) of a homogeneous more electron dense single layer, and a zona radiata interna, which comprised the innermost zona radiata (Z₃) of heterogeneous electron-dense multilayers. Especially, the surface of the outermost zona radiata (Z₁) showed a morphological difference in its external appearance among the species. In the surface structures of Z₁, I. yongdokensis had a round granule appearance, and both I. hugowolfeldi and I. longicorpus were villus in morphology. In the villus appearance of these two species, however, there were clear differences between I. hugowolfeldi and I. longicorpus: the former had a few short thin villi, 1.0–1.7 μm in length, but in the latter species the villi were numerous, long, and thick, 2.0–2.5 μm long. The variety of such structures in these three species of Iksookimia may be a useful aid in taxonomic relationships and closely related to their habitats. Received: February 15, 2000 / Revised: August 3, 2000 / Accepted: September 3, 2000     

Cytogenetic characterization of F1, F2 and backcross hybrids of the Neotropical catfish species Pseudoplatystoma corruscans and Pseudoplatystoma reticulatum (Pimelodidae, Siluriformes)

The cytogenetic characteristics of Pseudoplatystoma corruscans and Pseudoplatystoma reticulatum and their F1, F2 and backcross hybrids were assessed by using chromosome banding techniques. The diploid number of 56 chromosomes was constant in all species and lineages, with a karyotypic formula containing 20 metacentric, 12 submetacentric, 12 subtelocentric and 12 acrocentric chromosomes. Nucleolar organizer regions (NORs) were identified in two subtelocentric chromosomes in the parents and hybrids, with partial nucleolar dominance in F1 and F2 specimens. Heterochromatic blocks were detected in the terminal and centromeric regions of some chromosomes in all individuals. For parental and hybrid lineages, 18S ribosomal clusters corresponding to NORs and 5S ribosomal genes were identified in distinct pairs of chromosomes. The striking conservation in the chromosomal macrostructure of the parental species may account for the fertility of their F1 hybrids. Similarly, the lack of marked alterations in the chromosomal structure of the F1 hybrids could account for the maintenance of these features in post-F1 lineages.     

Patterns of reproduction and growth of the catfish Iheringichthys labrosus (Lütken, 1874) after a reservoir formation

This work contributes to the knowledge of the reproduction, growth parameters and mortality rates of Iheringichthys labrosus in a newly‐formed reservoir. A total of 554 males and 1227 females were collected over 12 consecutive months, 1998–1999, from sites in the Corumbá Reservoir, Brazil, using gillnets (meshes: 2.4–16 cm). Information on each individual, i.e. standard length (cm), weight (g), sex, and gonadal development phase was recorded. The pectoral spines were removed to estimate age. The number of juveniles and adults, males and females, reproductive sites and seasons were estimated. First maturation length was estimated using a likelihood function fitted by binomial distribution. Growth parameters were estimated using the von Bertalanffy equation. Total instantaneous mortality was obtained through a linearized catch curve method. Standard length varied from 6.0 to 20.5 cm. Growth showed negative allometry for both sexes. The reproduction period was August to December in all environments sampled and first maturation length was 11.5 cm. All individuals were adults with 17.0 cm standard length. Ages varied from zero to 7 years. Asymptotic length, growth coefficient and t₀ for the entire population were 27.79 cm, 0.12 and ?2.64, respectively. Instantaneous and annual mortality rates were 0.90 and 0.59, respectively.     

Seasonal growth and life history of the catfish Calophysus macropterus (Lichtenstein, 1819) (Siluriformes: Pimelodidae) from the Amazon floodplain

The objective was to study seasonal aspects of the piracatinga catfish ( Calophysus macropterus ) growth and its relationship with the flood pulse. Analyzed from June 1997 to May 1998 were 446 specimens from the Amazon River. Using the period regression model with several biometric indexes gave the following results: two periods of low growth were identified in the vertebrae of C. macropterus , the first during falling water (August) and the second during rising water (January), using marginal increment analysis (MIA). This pattern was also observed in the annual variation of the regression coefficient of the length–weight relationship ( b ), and Feeding Index (FI), but an inverse relationship was observed with the Maturity Index (MI). These results suggest that reproduction occurs from January to February in the rising water season, when C . macropterus reaches sexual maturity at 1.4 and 1.5 of age for females and males, respectively. The parameters of Bertalanffy's model were SL = 40.05[1−e^{−0.44(t−0.37)}] for females and SL = 41.9[1−e^{−0.42(t−0.36)}] for males. Natural mortality was high (♀:0.42–♂:0.40) and longevity low (♀:6.8–♂:7.1 years). The present study demonstrates that seasonal growth in C. macropterus is correlated with the flood pulse, especially in periods of rising and falling waters. These results, analyzed by the periodic regression model, showed that reproduction, migration or changes in feeding behavior have relationships with annulus formation in hard structures in this species. All quantitative indexes show clear seasonal variation to be correlated with river water level fluctuations.     

Genetic characterization of the Neotropical catfish Pimelodus maculatus (Pimelodidae,Siluriformes) in the Upper Uruguay River

Freshwater fish present unique challenges when one attempts to understand the factors that determine the structure of their populations. Habitat fragmentation is a leading cause of population decline that threatens ecosystems worldwide. In this study, we investigated the conservation status of genetic variability in the Neotropical catfish (Pimelodus maculatus). Specifically, we examined the structure and genetic diversity of this species in a region of the Upper Uruguay River fragmented by natural barriers and dams. There was no genetic structure among the four sites analyzed, indicating the existence of only one population group. A combination of environmental management and genetic monitoring should be used to minimize the impact of impoundment on panmitic populations of migratory fish species.     

The ultrastructure of Sorubim lima (Teleostei, Siluriformes, Pimelodidae) spermatogenesis: premeiotic and meiotic periods

The ultrastructure of Sorubim lima spermatogenesis during the premeiotic and meiotic periods was studied. Our observations showed that the germ cells in the cysts are connected by cytoplasmic bridges and the mitotic and meiotic divisions are slightly asynchronous. The first and the last spermatogonial generations differ in the cellular and nuclear volume, nucleolus, chromatin condensation, distribution, size, density, and shape of the mitochondria, presence of 'lamellae anulata', amount and dimension of the 'nuages', and movement of the centrioles. In addition to the nuclear prophase structures, the spermatocyte I shows changes in all other cellular organelles and elongated vesicles appear in the cytoplasm. The accentuated cytoplasmic density and thickened walled vesicles are morphological characteristics that differentiate spermatocytes II from the other germ cells in the cysts of Sorubim lima testis.     

Differences between the pathogenic processes induced by Steinernema and Heterorhabditis (Nemata: Rhabditida) in Pseudaletia unipuncta (Insecta: Lepidoptera)

Larvae of Pseudaletia unipuncta are moderately susceptible to infections caused by entomopathogenic nematodes, being a desirable host to study pathogenic processes caused by Heterorhabditis bacteriophora, Steinernema carpocapsae, and Steinernema glaseri and their associated bacteria. The ability of the infective stage of these nematodes to invade hosts is quite different. S. carpocapsae invades the highest number of insects and presents the highest penetration rate, followed by H. bacteriophora. Regression analysis between the number of insects parasitized and the number of IJs counted per insect, over time, showed a high correlation for S. carpocapsae whereas for H. bacteriophora it was low. Dose-response was most evident at a concentration below 100 IJs per insect on H. bacteriophora, whereas on S. carpocapsae it was found for doses ranging from 100 to 2,000 IJs. Student's t test analysis of dose-response showed parallel, yet unequal, slopes for both strains of H. bacteriophora, whereas distinct regressions were obtained for S. carpocapsae and S. glaseri, thus, evidencing each species develop a distinct pathogenic process. Insects injected with Photorhabdus luminescens died within 50 h after injection, whereas those treated with X. nematophila died much later. Moreover, the mortality in insects exposed to H. bacteriophora complex and injected with P. luminescens was close, but insects injected with bacteria died faster. Insect mortality in treatments with complexes S. carpocapsae and S. glaseri was significantly higher than that which was observed in insects injected with symbiotic bacteria.     

Virulence of Hypocreales fungi to pecan aphids (Hemiptera: Aphididae) in the laboratory

There is need for efficacious biocontrol agents for aphids in commercial orchards. As a preliminary step to this end we determined the virulence of several Hypocreales fungi to pecan aphids. In the first experiment we tested the virulence of Isaria fumosorosea (ARSEF 3581) blastospores to three pecan aphids Monellia caryella, Melanocallis caryaefoliae, and Monelliopsis pecanis under laboratory conditions. Rates of 1 × 10⁷ or 1 × 10⁸ spores per ml were applied in 2 ml via a spray tower to 90 mm Petri dishes containing 10 aphids each. Mortality and mycosis were determined after 24, 48 and 72 h. Treatment effects were observed by 48 h post-application, and by 72 h the higher application rate caused >90% mortality and mycosis in M. caryella and M. caryaefoliae, whereas <70% was observed in M. pecanis.We conducted two subsequent experiments (Experiments 2 and 3), using the same methodology, to compare the virulence of several Hypocreales species and strains against the aphid of primary economic concern to most pecan growers, M. caryaefoliae. In Experiment 2, we compared blastospores and conidia of two I. fumosorosea strains (ARSEF 3581 and ATCC 20874 [= strain 97]). The blastospores of ARSEF 3581 and conidia of ATCC 20874 showed higher virulence than other treatments and thus were included in Experiment 3, which also compared the virulence of conidia of Beauveria bassiana (GHA strain) and Metarhizium anisopliae (F52 strain). Results in Experiment 3 indicated the highest virulence in I. fumosorosea 3581 blastospores and M. anisopliae (F52) followed by I. fumosorosea (20874) conidia. The detection of pathogenicity to pecan aphids establishes the potential for commercial usage and additional study. Results reported here will narrow treatments to test in future greenhouse and field trials.     

Epizootiological studies of Amblyospora camposi (Microsporidia: Amblyosporidae) in Culex renatoi (Diptera: Culicidae) and Paracyclops fimbriatus fimbriatus (Copepoda: Cyclopidae) in a bromeliad habitat

The epizootiology of Amblyospora camposi was studied in a natural population of Culex renatoi, a bromeliad-inhabiting mosquito, and its intermediate host, Paracyclops fimbriatus fimbriatus, over a 2-year period. Twenty Eryngium cabrerae plants were sampled monthly from January 2003 to January 2005 and the prevalence of A. camposi in P.f. fimbriatus and Cx. renatoi populations was determined. The monthly prevalence rates of meiospore infections in Cx. renatoi larvae never exceeded 5.5% and was detected in 50% of the monthly samples. Meiospores were available in plants over the course of the study at a mean concentration of 2 x 10(4) meiospores/ml. Within each plant the parasite was maintained by horizontal transmission. P.f. fimbriatus with vegetative stages and mature spores were found regularly in bromeliads suggesting efficient meiospore infectivity to field copepod populations. The mean concentration of spores from copepods found in plants was 8 x 10(2) spores/ml. Infections in copepods were detected in 54% of the monthly samples with a prevalence rate ranging from 0.55 to 17.4% and an overall average of 5.1%. Vegetative stages in fourth instar mosquito larvae (probably derived from the horizontal pathway via spores formed in copepods) were detected in 12.5% of the monthly samples with an overall prevalence rate of 1.1%. Infections in female and male adults were detected in 20.8% of the monthly samples with an overall average of 4.1% and 6.8%, respectively.     

Serous goblet cells: The protein secreting cells in the oral cavity of a catfish, Rita rita (Hamilton, 1822) (Bagridae,Siluriformes)

Serous goblet cells in the oral epithelium of Rita rita are characterized by the presence of distinct eosinophilic granules occupying large parts of the cytoplasm. In R. rita, a range of histochemical results reveal that these cells are involved in proteinaceous secretions, and thus likely contribute to various functions analogous to those of mammalian saliva. The secretions of these cells have also been associated with specific functions and are discussed in relation to their physiological importance with special reference to their roles in lubrication, alteration in viscosity, various functions of mucus such as handling, maneuvering and driving of food items toward the esophagus, maintaining taste sensitivity and protection of the oral epithelium. In addition, the serous goblet cells may also be considered as the primary defensive cell of the oral epithelium of R. rita. The results significantly add to very limited set of literature on the serous goblet cells and provide noteworthy information on the mucous secretions in the oral cavity of fish.     

Evolutionary history of frogfishes (Teleostei: Lophiiformes: Antennariidae): a molecular approach

Fishes of the family Antennariidae (order Lophiiformes) are primarily shallow-water benthic forms found in nearly all tropical and subtropical oceans and seas of the world, with some taxa extending into temperate waters. Despite an earlier attempt based on morphology, no previous hypothesis of intergeneric relationships of the Antennariidae exists. To resolve phylogenetic relationships within the Antennariidae, and to test the validity of species groups within Antennarius, DNA sequences from the mitochondrial 16S and cytochrome oxidase c subunit 1 (COI) genes, and nuclear recombination activating gene 2 (RAG2), for 25 described and four undescribed antennariid species, representing 10 of 12 known genera and one undescribed genus, were unambiguously aligned and analyzed using Bayesian and maximum likelihood methods. The markers were partitioned and analyzed for substitution saturation and only the third codon position of COI (COI-3) was found to have reached saturation. However, analysis of both datasets, one with the saturated data and one without, differed only slightly. All molecular analyses recovered two major clades, one comprised of Fowlerichthys, Antennarius, Histrio, and Antennatus; and another containing Rhycherus, Antennariidae gen. et sp. nov., Kuiterichthys, Phyllophryne, Echinophryne, Tathicarpus, Lophiocharon, and Histiophryne. Evidence is presented to illustrate a correlation between phylogeny, geographic distribution, and reproductive life history. The results of these analyses provide the first hypothesis of evolutionary relationships within the Antennariidae.     

Localization and enzymatic activity profiles of the proteases responsible for tachykinin-directed oocyte growth in the protochordate, Ciona intestinalis

We previously substantiated that Ci-TK, a tachykinin of the protochordate, Ciona intestinalis (Ci), triggered oocyte growth from the vitellogenic stage (stage II) to the post-vitellogenic stage (stage III) via up-regulation of the gene expression and enzymatic activity of the proteases: cathepsin D, carboxypeptidase B1, and chymotrypsin. In the present study, we have elucidated the localization, gene expression and activation profile of these proteases. In situ hybridization showed that the Ci-cathepsin D mRNA was present exclusively in test cells of the stage II oocytes, whereas the Ci-carboxypeptidase B1 and Ci-chymotrypsin mRNAs were detected in follicular cells of the stage II and stage III oocytes. Double-immunostaining demonstrated that the immunoreactivity of Ci-cathepsin D was largely colocalized with that of the receptor of Ci-TK, Ci-TK-R, in test cells of the stage II oocytes. Ci-cathepsin D gene expression was detected at 2h after treatment with Ci-TK, and elevated for up to 5h, and then slightly decreased. Gene expression of Ci-carboxypeptidase B1 and Ci-chymotrypsin was observed at 5h after treatment with Ci-TK, and then decreased. The enzymatic activities of Ci-cathepsin D, Ci-carboxypeptidase B1, and Ci-chymotrypsin showed similar alterations with 1-h lags. These gene expression and protease activity profiles verified that Ci-cathepsin D is initially activated, which is followed by the activation of Ci-carboxypeptidase B1 and Ci-chymotrypsin. Collectively, the present data suggest that Ci-TK directly induces Ci-cahtepsin D in test cells expressing Ci-TK receptor, leading to the secondary activation of Ci-chymotrypsin and Ci-carboxypeptidase B1 in the follicle in the tachykininergic oocyte growth pathway.     

Phylogenetic relationships among Southeast Asian climbing bamboos (Poaceae: Bambusoideae) and the Bambusa complex

A phylogenetic analysis of Bambusa and allies based on the plastid DNA non-coding regions rps16-trnQ, trnC-rpoB, trnH-psbA and trnD-T, and a partial nuclear GBSSI gene, was carried out. This included representatives from all four Bambusa subgenera (including type species), a group of segregate Southeast Asian genera distinctive by their climbing–scrambling culms (Dinochloa, Holttumochloa, Kinabaluchloa, Maclurochloa, Soejatmia, Sphaerobambos), and two other Bambusinae genera (Dendrocalamus, Gigantochloa). The results do not support the present subgeneric classification of Bambusa. The climbing Southeast Asian genera, all of which include species previously placed in Bambusa, are distinct from the “core Bambusa group” (type species and alliance) and the Bambusa complex generally.     

Microscopic and molecular characterization of ovarian follicle atresia in Rhodnius prolixus Stahl under immune challenge

In this work we characterized the degenerative process of ovarian follicles of the bug Rhodnius prolixus challenged with the non-entomopathogenic fungus Aspergillus niger. An injection of A. niger conidia directly into the hemocoel of adult R. prolixus females at the onset of vitellogenesis caused no effect on host lifespan but elicited a net reduction in egg batch size. Direct inspection of ovaries from the mycosed insects revealed that fungal challenge led to atresia of the vitellogenic follicles. Light microscopy and DAPI staining showed follicle shrinkage, ooplasm alteration and disorganization of the monolayer of follicle cells in the atretic follicles. Transmission electron microscopy of thin sections of follicle epithelium also showed nuclei with condensed chromatin, electron dense mitochondria and large autophagic vacuoles. Occurrence of apoptosis of follicle cells in these follicles was visualized by TUNEL labeling. Resorption of the yolk involved an increase in protease activities (aspartyl and cysteinyl proteases) which were associated with precocious acidification of yolk granules and degradation of yolk protein content. The role of follicle atresia in nonspecific host-pathogen associations and the origin of protease activity that led to yolk resorption are discussed.     

Ultrastructural study of spermiogenesis and the spermatozoon of the proteocephalidean cestode Barsonella lafoni de Chambrier et al., 2009, a parasite of the catfish Clarias gariepinus (Burchell, 1822) (Siluriformes, Clariidae)

Spermiogenesis in the proteocephalidean cestode Barsonella lafoni de Chambrier et al., 2009 shows typical characteristics of the type I spermiogenesis. These include the formation of distal cytoplasmic protrusions forming the differentiation zones, lined by cortical microtubules and containing two centrioles. An electron-dense material is present in the apical region of the differentiation zone during the early stages of spermiogenesis. Each centriole is associated to a striated rootlet, being separated by an intercentriolar body. Two free and unequal flagella originate from the centrioles and develop on the lateral sides of the differentiation zone. A median cytoplasmic process is formed between the flagella. Later these flagella rotate, become parallel to the median cytoplasmic process and finally fuse proximodistally with the latter. It is interesting to note that both flagellar growth and rotation are asynchronous. Later, the nucleus enlarges and penetrates into the spermatid body. Finally, the ring of arching membranes is strangled and the young spermatozoon is detached from the residual cytoplasm.The mature spermatozoon presents two axonemes of the 9 + ‘1’ trepaxonematan pattern, crested body, parallel nucleus and cortical microtubules, and glycogen granules. Thus, it corresponds to the type II spermatozoon, described in almost all Proteocephalidea. The anterior extremity of the gamete is characterized by the presence of an apical cone surrounded by the lateral projections of the crested body. An arc formed by some thick and parallel cortical microtubules appears at the level of the centriole. They surround the centriole and later the first axoneme. This arc of electron-dense microtubules disorganizes when the second axoneme appears, and then two parallel rows of thin cortical microtubules are observed. The posterior extremity of the male gamete exhibits some cortical microtubules. This type of posterior extremity has never been described in proteocephalidean cestodes. The ultrastructural features of the spermatozoon/spermiogenesis of the Proteocephalidea species are analyzed and compared.     

Genetic structure and historical diversification of catfish Brachyplatystoma platynemum (Siluriformes: Pimelodidae) in the Amazon basin with implications for its conservation

Brachyplatystoma platynemum is a catfish species widely distributed in the Amazon basin. Despite being considered of little commercial interest, the decline in other fish populations has contributed to the increase in the catches of this species. The structure, population genetic variability, and evolutionary process that have driven the diversification of this species are presently unknown. Considering that, in order to better understand the genetic structure of this species, we analyzed individuals from seven locations of the Amazon basin using eight molecular markers: control region and cytochrome b mtDNA sequences, and a set of six nuclear microsatellite loci. The results show high levels of haplotype diversity and point to the occurrence of two structured populations (Amazon River and the Madeira River) with high values for F_ST. Divergence time estimates based on mtDNA indicated that these populations diverged about 1.0 Mya (0.2–2.5 Mya 95% HPD) using cytochrome b and 1.4 Mya (0.2–2.7 Mya 95% HPD) using control region. During that time, the influence of climate changes and hydrological events such as sea level oscillations and drainage isolation as a result of geological processes in the Pleistocene may have contributed to the current structure of B. platynemum populations, as well as of differences in water chemistry in Madeira River. The strong genetic structure and the time of genetic divergence estimated for the groups may indicate the existence of strong structure populations of B. platynemum in the Amazon basin.     

Dietary nitrogen availability in macroalgae enhances growth of the sea hare Aplysia californica (Opisthobranchia: Anaspidea)

Evidence suggests that marine herbivores select for prey items with elevated nitrogen content. We tested this hypothesis with experimental growth studies of the herbivorous gastropod Aplysia californica offered diets of the rhodophyte Gracilaria ferox with varying nitrogen content. A. californica had a sevenfold feeding preference by weight for G. ferox with higher tissue % N and % protein, and lower % C, % carbohydrate and carbon-to-nitrogen ratio (C/N) as compared to treatments with lower nitrogen and protein content. A. californica provided treatment diets of varying nitrogen availability (high, medium and low) also had significantly different growth rates within 30 days. The high N diet supported a two-fold higher growth rate than that of the medium N diet and four-fold higher than the low N treatment, with mean weights over the experiment reaching 20.4, 9.8, and 5.6 g, respectively. The evolutionary value of this dietary preference may facilitate faster growth to reproductive size, outgrowth of predators and sequestration of organic compounds from nitrogen-enriched macroalgae to deter predators. Despite recent attention, the importance of diet-derived compounds as anti-predatory chemical defenses in Aplysiids remains equivocal. Nitrogen-enriched macroalgal diets, however, may provide “primary metabolites” that serve as alternative defenses for Aplysiids.     

Influence of host plants on feeding, growth and reproduction of Papilio polytes (The common mormon)

We studied the feeding, growth and reproductive behaviour of Papilio polytes (common mormon butterfly) on five different host plants, Murraya koenigii, Toddalia asiatica, Glycosmis pentaphylla, Aegle marmelos and Citrus medica. The growth rate of P. polytes was fastest on M. koenigii followed by T. asiatica, C. medica, G. pentaphylla and A. marmelos. We related this to the nutrient contexts of the five plants. The plants T. asiatica and C. medica had higher water contents, which influenced the growth rate of the insect. M. koenigii was found to contain rich quantities of carbohydrate. M. koenigii, T. asiatica and C. medica were also rich in protein when compared to A. marmelos and G. pentaphylla. Total amino acid levels were comparatively higher in M. koenigii, T. asiatica, C. medica rather than A. marmelos and G. pentaphylla.