Puntius Conchonius Research Articles

Lipovitellin is a Non-Self Recognition Receptor with Opsonic Activity

Lipovitellin (Lv), a glycolipoprotein, is a major component of the egg yolk, which is usually regarded as an energy reserve of nutrients essential for growth and development. We have purified Lv from ovulated eggs of the rosy barb Puntius conchonius by two-step chromatography and characterized it by staining with periodic acid/Schiff reagent and Sudan black B, amino acid composition analysis, and peptide mass fingerprinting. The results of ligand and bacterial binding assays, an enzyme-linked immunosorbent assay (ELISA), and the phagocytosis test revealed, for the first time, that the purified native form of P. conchonius Lv acts as a pattern recognition molecule with multiple specificities capable of identifying pathogen-associated molecular patterns (PAMPs), including those of lipopolysaccharide, lipoteichoic acid, and peptidoglycan, rather than self components and that it can bind Gram-negative and -positive bacteria, such as Escherichia coli and Staphylococcus aureus. These tests also showed that the P. conchonius Lv functions as an opsonin capable of enhancing macrophage phagocytosis. Taken together, these characteristics suggest that in developing embryos/larvae of P. conchonius, the native form of Lv may be physiologically involved in the sensing of invading pathogens via interaction with PAMPs and in the recruitment of the primitive macrophages that appear in early embryos to phagocytose and digest the pathogens, thereby protecting them from pathogenic attacks.

Fright reaction in light and dark: how visual information availability modulates the response to chemical alarm cues

Alarm cues, released when the skin of ostariophysan fish is injured, elicit striking anti-predator behaviour in conspecifics. Recent evidence suggests that environmental conditions may affect the response. Light and visibility, and thus the possibility of using the visual sensory channel, may greatly vary in aquatic habitats. The aim of this study was to evaluate the role of visual stimuli in the expression of the response to an alarm cue by Barbus conchonius Hamilton–Buchanan 1822, by analysing both swimming patterns and spatial distribution of the fish. Fish were tested in light or darkness in the absence or presence of a chemical alarm cue. In the absence of light, a simplified fish behavioural response ensued to an alarm cue, i.e., the typical swimming patterns were not performed. However, alteration of fish spatial distribution caused by alarm cue perception was unaffected by the impossibility of relying on visual cues, as it also occurred in the dark. An increase in cohesion and downward movement in the dark may have an adaptive value in such an environmental condition. However, we hypothesise that swimming pattern alteration does not occur in the dark because it has evolved as a visual signal for conspecifics and/or as an antipredatory response specific for visual hunting predators.

Changes in extracellular osmolality initiate sperm motility in freshwater teleost rosy barb Puntius conchonius

The objective was to investigate the effects of extracellular osmolality and membrane osmotic-sensitive channels on the initiation of sperm motility and to explore mechanisms of sperm initiation in rosy barb ( Puntius conchonius). We found that (1) sperm were immotile in seminal plasma and remained quiescent in electrolyte or nonelectrolyte solutions isotonic to seminal plasma; (2) sperm movement was initiated when the sperm were exposed to hypo-osmotic electrolyte or hypo-osmotic nonelectrolyte solutions, and that the responsiveness of sperm to changes in the extracellular osmolalities (100, 200, 250, 270, and 300 mOsm/kg) differed among sperm cells (P < 0.05); (3) sperm movement could be initiated and terminated repeatedly by decreasing and increasing the osmolality (in increments of 100 and 300 mOsm/kg) of a nonelectrolyte mannitol solution, respectively (P < 0.05); (4) gadolinium (20, 40, and 80 μM) inhibited the initiation of sperm motility and abolished the sperm activation caused by the hypo-osmotic media treatment in dose- and time-dependent manners (P < 0.05); and (5) sperm activation in a hypo-osmotic medium and inhibition in an isotonic solution were associated with swelling and shrinkage of the sperm sleeves, respectively. Therefore, we concluded that osmolality was a critical physiologic signal in regulating the initiation and termination of sperm motility in freshwater teleost rosy barb. Furthermore, we inferred that rosy barb sperm were hypo-osmotic–dependent conformers, and the osmotic-sensitive channel could be involved in the mechanism of sperm initiation.

Corrigendum

This article refers to:Comparison of Histopathological Alterations Due to Sublethal CCl4 on Rosy Barb (Puntius conchonius) and Amphioxus (Branchiostoma belcheri) with Implications of Liver Ontogeny

Toxicity studies of nonylphenol on rosy barb ( Puntius conchonious): A biochemical and histopathological evaluation

Acute and subacute toxicity of the nonylphenol (NP) on fish was investigated in laboratory toxicity tests with rosy barb ( Puntius conchonious). The acute toxicity of NP to rosy barb was determined in semi-static bioassays. Median lethal concentration (LC 50) for 96 h was 1.72 μM. The effects of sublethal concentrations of NP (0.17, 0.34 and 0.68 μM) on the structures and biochemical parameters [alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alanin aminotransferase (ALT)] in gills, liver and kidney of rosy barb were studied after 14 days. The results showed that NP caused alteration of the structure in organs, as evidenced by the hyperplasia of epithelium and the fusion of secondary lamellae in the gills, the disappearance of the cell membrane and the cell necrosis in the liver as well as haemorrhages in the kidney. In addition, the functional enzyme activities were also changed. The increase trend in ALP activity in organs of fish treated with NP was recorded. The levels of AST and ALT in gills, liver and kidney were stimulated to rise at the lower concentration and fall at the higher concentration NP treatment compared to controls. This study suggests that NP can alter of the structures and biochemical parameters within non-endocrine tissue of fish and these changes may be mediated via destroying membrane structure and inducing cell necrosis.

Comparison of Histopathological Alterations Due to Sublethal CCl4 on Rosy Barb (Puntius conchonius) and Amphioxus (Branchiostoma belcheri) with Implications of Liver Ontogeny

ABSTRACTThis study was undertaken to examine the histopathological effects of CCl4 on rosy barbs and amphioxus with an aim to compare the homology between rosy barb liver and amphioxus digestive caecum. It was found that the 96 h LC50 values were 23.9 ± 4 mg/L and 18.9 ± 2 mg/L for rosy barbs and amphioxus, respectively. Histological examinations showed that exposure to sublethal CCl4 caused damage to the liver, kidney, and gill in rosy barb, and to the digestive caecum and gill in amphioxus. It is clear that both rosy barb liver and amphioxus digestive caecum were the prominent target organs of CCl4, suggesting that the digestive caecum in amphioxus is homologous to the liver in rosy barb at least in respect to toxic damages of CCl4.

Early gonadal development and sex differentiation in rosy barb (Puntius conchonius)

AbstractThe main objective of this study was to describe the early gonadal development and to examine the process of sex differentiation in male and female P. conchonius under laboratory conditions. First evidence of primordial germ cells was observed on the day of hatching. The sex differentiation in leptotene, zygotene, pachytene and diplotene stages was clearly detected. Differentiation started from the mid-mid posterior part of the gonads. Actual sex differentiation occurred between 18-21 days and 36-40 days post-hatching in females and males, respectively. Histological sex differentiation differences were clear between males and females; in males, gonads had a smooth surface, were less stained, arrow-shaped, with germ cells located alone in the stroma, and number of germ cells ranging from two to ten per section, whereas in females, gonads had a rough surface, were more stained, wider, with germ cells multiplying rapidly and forming clusters, and number of germ cells ranged from 2 to 58 per section. The numbers of germ cells within the two gonad types were significantly different in favour of females (P &lt; 0.05). Here early developmental stages of the gonads from 0 to 56 days post-hatch are explained.

Vitellogenin is a novel player in defense reactions

The significance of the presence of vitellogenin (Vg), a female-specific protein, in male and juvenile fish is uncertain. Here we show that the Vg purified from the rosy barb Puntius conchonius possesses both antibacterial and hemagglutinating activities in vitro, and the male fish challenged with Escherichia coli synthesize Vg. These suggest that in addition to being involved in yolk protein formation, Vg is also involved in defense reactions, a novel function assigned to Vg.

Embryonic development of the rosy barbPuntius conchoniusHamilton 1822 (Cyprinidae)

This study provides the first detailed table of normal development of the rosy barb embryo, from fertilization to post-hatching. We define six broad periods of embryogenesis, i.e. the zygote, cleavage, blastula, gastrula, segmentation and larval periods. These divisions highlight the changing spectrum of major developmental processes occurring during these periods. Stages that subdivide the periods are named, not numbered as in most other series. The stages and their names are based on morphological features usually readily identified by examination of the live embryo under a stereomicroscope. These provide a detailed embryological staging by morphological criteria for future studies on molecular biology and biotechnology in this species.

Genetic analysis of offspring from intra‐ and interspecific crosses of Carassius auratus gibelio by chromosome and RAPD analysis

The ploidy of silver crucian carp Carassius auratus gibelio individuals, originating from nine natural habitats of Hungary, was estimated by erythrocyte nucleus area analysis. On the basis of DNA polymorphism, the genetic homogeneity or heterogeneity and the chromosome number of different offspring derived from the crossing of triploid and diploid populations and of two types of silver crucian carp females with other cyprinid males (Cyprinus carpio, Carassius carassius, Carassius auratus and Barbus conchonius) were determined. The results of chromosome and RAPD analysis demonstrated that diploid females could reproduce sexually with silver crucian carp and other cyprinid males and that the offspring of intra‐ and interspecific crosses contained the paternal DNA. Triploid females usually reproduced by gynogenesis and their offspring were clones, however, in very rare cases paternal genes were actually transmitted (i.e. paternal leakage) to the offspring and the progeny were triploid interspecific hybrids. RAPD analysis showed that while the paternal DNA appeared in the offspring, the maternal phenotype was strongly expressed.

Molecular and morphological data suggest that Spinibarbus caldwelli (Nichols) (Teleostei: Cyprinidae) is a valid species

The taxonomic problem of the cyprinid species of genus Spinibarbus, occurring in southern China and northern Vietnam, was resolved on the basis of molecular and morphological analyses. Spinibarbus caldwelli and Spinibarbus hollandi have a smooth posterior edge of the last unbranched dorsal fin ray among species in the genus. Spinibarbus caldwelli is currently regarded as a junior synonym of S. hollandi because of ambiguities in diagnostic characters. In this article, 11 mtDNA cytochrome b sequences of Spinibarbus specimens were analyzed together with Barbodes gonionotus and Puntius conchonius as outgroups. Our results showed that specimens identified as S. hollandi from Taiwan were different from those from the Asian mainland at a high level of genetic divergence (0.097-0.112), which is higher than that between the two valid species, S. sinensis and S. yunnanensis ( 0.089), and suggested that Taiwan specimens should be considered as a different species from the Asian mainland one. In a molecular phylogenetic analysis, the sister-group relationship between Taiwan specimens and the Asian mainland specimens was supported strongly by a high confidence level ( 100% in bootstrap value). Further analysis of morphological characters showed that overlap of diagnostic characters is much weaker than previously suggested. Taiwan specimens had 8 branched rays in the dorsal fin, whereas those from the mainland had almost 9-10. The molecular and morphological differences suggest S. caldwelli to be valid. The molecular divergence shows the genetic speciation of S. hollandi and S. caldwelli might have occurred 5.6-4.9 million years ago; the former could be a relict species in Taiwan, and the latter dispersed in the Asian mainland.

Exposure of Rosy Barb (Puntius conchonius) Sperm to Abamectin as an In Vitro Assay of Cytotoxicity

The acute cytotoxicity of abamectin to the sperm of rosy barb (Puntius conchonius) was tested. It was found that 79% sperm remained motile at 30 min after activation by hypo-osmotic sodium buffer (HSB), while the motility of sperm exposed to 0.14, 0.28, and 0.42 μ M abamectin for 30 min was reduced to 60%, 35%, and 15%, respectively. Exposure to abamectin at 0.56 μ M decreased the motility of sperm to an even lower value of 5.67% after 20 min of exposure. This indicates that the toxicity of abamectin to rosy barb sperm is both dose dependent and exposure time dependent. The ultrastructural examination showed that most of the sperm exposed to 0.14 μ M abamectin for 30 min remained morphologically normal, while the midpiece and its surrounding mitochondria of the sperm treated with 0.28 μ M abamectin for 30 min were severely damaged. For the sperm treated with 0.42 μ M abamectin for 30 min, the majority of the sperm were abnormal. Sperm heads were dissolved and the midpiece broken. The mitochondria are located in the midpiece of the sperm and may be the target of abamectin toxicity in rosy barb sperm. The results also suggest that rosy barb sperm are a suitable candidate for rapid screening of the acute toxicities of bioorganic pesticides.

Production and progeny testing of androgenetic rosy barb Puntius conchonius

Protocol for androgenetic cloning of the rosy barb, Puntius conchonius, with contrasting gray and golden strains is described. At the intensity of 4.2 W/m2, UV irradiation for 3.0 min inactivates the maternal genome in eggs of the gray barb. Following activation by the golden barb sperm, 24-min old eggs are shocked at 41 degrees C for 2 min to restore diploidy. Maternal genomic inactivation is confirmed by the (i) golden body color, (ii) karyotyping, and (iii) progeny testing of F1-F3 progenies. Estimates of stage-specific mortality of haploid and diploid androgenotes indicate no change in the time scale or developmental sequence, when sperm of related strain is used for activation, and when haploid genome regulates the development. Survival of androgenetic clones remains constant for the F1, F2, and F3 progenies and is about 15% and 7% at hatching and sexual maturity, respectively. Homozygosity of the androgenotes is shown to inflict greater mortality. Between F1 and F3 generations, the heterozygosity of the androgenetic clone is decreased, as evidenced by reduction in size hierarchy. Though the reproductive performance of the F1, F2, and F3 supermales is superior to the normal ones, the realized fecundity remains equal around 80 progenies per brood. The 92 crosses involving 16 supermales and 10 normal dams yield 75-100% male progenies, confirming the possible operation of XXfemale symbol:XYmale symbol sex determination system. The frequency of unexpected occurrence of female progenies is about 8%, the causes for which are discussed.

Use of heterologous sperm for the dispermic induction of androgenesis in barbs

Dispermic activation of genome‐inactivated eggs of the grey tiger barb Puntius tetrazona, facilitated by 2·5% polyethylene glycol (PEG)‐incubated sperm of the golden rosy barb Puntius conchonius, resulted in the generation of interspecific androgenetic clones of the golden rosy barb. A 10 min incubation of the golden rosy barb sperm in increasing concentrations (1·0, 1·5, 2·0, 2·5 and 3·0%) reduced the frequency of motile sperm (to 70%), motility duration (110–50 s) and fertilizability (to 80%) of the sperm; however, the frequency of sperm with double head size increased. At 3% PEG, motility pattern of the sperm completely changed from ‘zig‐zag’ to ‘circular’. Incubation in 2·5% PEG facilitated the dispermic entry and production of diploid androgenetic female and male progenies at the ratio of 0·27 : 0·73. Polymerase chain reaction (PCR) analysis using Tc1 transposon specific primers confirmed the purity of paternal inheritance of P. conchonius through the surrogate eggs of P. tetrazona. Survival and breeding, body colour, diploidy (karyotyping and erythrocyte measurements) and 0·27F : 0·73M sex ratio of the androgenotes provided evidence for the successful induction of dispermic androgenesis. Despite increased heterozygosity and reduced ‘damage cost’ on restoration of diploidy, survival of dispermic androgenotes induced in heterologous eggs was lower (1·7%) than those reported for androgenetic golden rosy barbs induced using homologous sperm (14·0%) and heterologous eggs (7·0%), i.e. tiger barb eggs.

Hormonal induction of supermale golden rosy barb and isolation of Y-chromosome specific markers

Viable supermales of the golden rosy barb Puntius conchonius are generated through hormonal sex reversal and progeny testing. Discrete immersion (3 h/day on the second, fourth, and sixth day after hatching) of the fry in Estradiol-17β (E 2) at doses of 400–600 μg/L ensures greater than 40% survival and production of more than 98% F 1 females; of these 50% are heterogametic females, that when crossed with normal males, sire 25% YY supermales. Supermales sire female progenies at the frequencies between 0 and 8%. Reproductive performance of hormonally sex-reversed females and androgenetic females is inferior to the normal ones. Conversely, the performance of androgenetic males is superior but suffers from low fertilizability. The relative performance of supermale produced by breeding sex-reversed parents is superior to those produced by androgenesis. Using the SRY-specific primers, the PCR analysis of the genomic DNA of the male golden rosy barb produces three products of 588, 333, and 200 bp length. However only the 200 bp product is amplified in the female genome. Hence it is possible to use the first two products as molecular markers to rapidly identify fish possessing a Y chromosome. The presence of 333 and 588 bp fragments in normal (X 1Y 2), hormonally induced (Y 1Y 2) and androgenetic (Y 2Y 2) males and the absence of relation between the 200 bp fragment and the X-chromosome indicates that the male specific markers are specific to Y-chromosome. For the first time, a Y-chromosome specific molecular marker for a cyprinid has been identified, isolated, and characterized.

Locomotory Responses of the Cave Cyprinid Phreatichthys Andruzzii Chemical Signals from Conspecifics and Related Species: New Findings

Previous research on chemical communication among hypogean fishes have suggested that these species are attracted, as are epigean species, by the odour of non-injured conspecifics. Considerations on the biological meaning of such responses and some doubts about the reliability of the experimental procedure previously utilised, have led us to reconsider the matter and verify the results already obtained in the cave cyprinid Phreatichthys andruzzii. Using a new methodological approach we ascertained not only the absence of positive chemotaxis to odour of known or unknown conspecifics, but also the presence of a strong negative chemotaxis towards the odour of unfamiliar conspecifics and of individuals belonging to the closely related species Barbus conchonius. The biological meaning and the adaptive value of these reactions are briefly discussed.

Fish Diversity of Kolli Hills, Western Ghats, Salem District, Tamil Nadu

Received on 24 April 2000 Accepted on 29 November 2000 Abstract The fish diversity of Periyaru originating as a natural spring in Kolli Hills is reported. A total number of 19 species of fish belonging to 15 genera and nine families were collected. The percentage occurrence is 36.88 for Cyprinidae, Bagridae and Cichlidae constituted each by 15.8. Of the 19 species, Ompok bimaculatus and Pseudeutropius atherinoides are endangered and Barilius bendelisis, Puntius sophore, Labeo calbasu and Mystus cavasius come under Low Risk nearly threatened (LRnt) category and Cirrhinus reba, Puntius conchonius, Mystus vittatus, Mystus bleekeri, Awaous gutum, Clarias batrachus and Mastacembelus armatus are vulnerable species.

Induction of diploid gynogenesis using interspecific sperm and production of tetraploids in African catfish, Clarias gariepinus Burchell (1822)

Homozygous diploid gynogenetic and tetraploid progeny of African catfish ( Clarias gariepinus) were produced by the suppression of the first mitotic division by applying thermal shock. The duration of one mitotic cycle during synchronous cell divisions ( τ 0-value) was used as unit after fertilization for expression of the time of development. After determination of sensitive and non-sensitive periods, a temperature of 40.5°C for 2 min was subsequently used. Diploid gynogenesis was obtained (6.34±2.35% survivors at swimming stage) when eggs were fertilized with genetically inactivated rosy barb ( Barbus conchonius) sperm. We have not found any embryos using intact interspecific sperm. When African catfish sperm was used for fertilization a ratio of 9.2% tetraploids was obtained (mixed eggs from four females). The highest frequency of both gynogens and tetraploids was observed when the heat shock was initiated at 1.4 τ 0-value. We used electrophoresis, chromosomes and nucleoli counting for identification of homozygosity and tetraploidy.

Production of fish chimeras from embryonic cells

The efficiency of two cell-transplantation methods were compared for the production of embryonic cell derived fish chimeras. The classic microinjection technique (blastula stage donor cells are microinjected into blastula embryos) was compared to a novel aggregation method for fish developed by our group. This method utilises the ability of dechorionated fish embryos to aggregate. Morula cells dissociated in Ca2+, Mg2+ free medium were aggregated to recipient embryos of different developmental stage. Donor cells and recipient embryos of different developmental stages were used for most efficient incorporation in the chimeras. THe fate of donor cells derived from blastulae, was followed by labelling them with FITC-dextrane (FD). The most efficient transplantations were gained by using 16-32 cell stage recipients for aggregation (18% survival of chimeras at swim up stage). Labelled donor cells were contributing to the embryos in varying ratio. A comparison of the efficiency of aggregation was made between diploid-diploid, diploid-haploid and diploid-interspecfic (diploid) hybrid chimeras. In all three cases chimeras containing different proportion of donor cells were gained. After one day incubation the embryos were dissociated by trypsin digestion and number of labelled and non-labelled cells were counted under fluorescent microscope. Experiments were performed on Rosy barb (Barbus conchonius), Carp (Cyprinus carpio) and African catfish (Clarias gariepinus).

Fungal diseases of fish in Nanak Sagar, Naini Tal, India.

Eight zoosporic fungi viz., Achlya debaryana, A. flagellata, A. klebsiana, Aphanomyces laevis, Saprolegnia diclina, S. ferax, S. parasitica, and Pythium sp. were isolated from a large number of adult fishes of the species Mastacembelus armatus, Mystus vitatus, Nandus nandus, Tor putitora and T. tor of Nanak Sagar reservoir in Naini Tal district, India. Species of the parasites and the hosts were different in their pathogenicity and immunity, respectively. However, A. flagellata and S. parasitica appeared to be the most virulent. The severity of mycosis was primarily correlated to moderate water temperatures of 22-25 degrees C. High temperature (> 28 degrees C) retarded the disease process. The experimental inoculation with all the associated fungal species on Puntius conchonius in the laboratory produced clinical signs similar to the ones seen on infected fish in the reservoir. This is the first report on fish mycosis in the large reservoir located in the foot hill of Kumaun Himalaya.