Genetic heterogeneity and ploidy level analysis among different gynogenetic clones of the polyploid gibel carp

Background: Some triploid and tetraploid clones have been identified in the gynogenetic gibel carp, Carassius auratus gibelio Bloch, by karyotypic and cytologic analyses over many years. Further, 5-20% males and karyotypic diversity have been found among their natural and artificial populations. However, the DNA contents and the relation to their ploidy level and chromosome numbers have not been ascertained, and whether normal meiosis occurs in spermatogenesis needs to be determined in the different clones. Methods: The sampled blood cells or sperms were mixed with blood cells from chicken or individual gibel carp and fixed in 70% pre-cooled ethanol overnight at 4degreesC. The mixed cell pellets were washed 2-3 times in 1x phosphate buffered saline and then resuspended in the solution containing 0.5% pepsin and 0.1 M HCl. DNA was stained with propidium iodide solution (40 mug/mL) containing 4 kU/ml RNase. The measurements of DNA contents were performed with Phoenix Flow Systems. Results: Triploid clones A, E, F, and P had almost equal DNA content, but triploid clone D had greater DNA content than did the other four triploid clones. DNA content of clone M (7.01 +/- 0.15 pg/nucleus) was almost equal to the DNA content of clone D (5-38 +/- 0.06 pg/nucleus) plus the DNA content of common carp sperm (1.64 +/- 0.02 pg/nucleus). The DNA contents of sperms from clones A, P, and D were half of their blood cells, suggesting that normal meiosis occurs in spermatogenesis. Conclusions: Flow cytometry is a powerful method to analyze genetic heterogeneity and ploidy level among different gynogenetic clones of polyploid gibel carp. Through this study, four questions have been answered. (a) The DNA content correlation among the five triploid clones and one multiple tetraploid clone was revealed in the gibel carp, and the contents increased with not only the ploidy level but also the chromosome number. (b) Mean DNA content was 0.052 pg in six extra chromosomes of clone D, which was higher than that of each chromosome in clones A, E, F, and P (about 0.032 pg/ chromosome). This means that the six extra chromosomes are larger chromosomes. (c) Normal meiosis occurred during spermatogenesis of the gibel carp, because DNA contents of the sperms from clones A, P, and D were almost half of that in their blood cells. (d) Multiple tetraploid clone M (7.01 +/- 0.15 pg/nucleus) contained the complete genome of clone D (5.38 +/- 0.06 pg/nucleus) and the genome of common carp sperm (1.64 +/- 0.02 pg/nucleus). Cytometry Part A 56A:46-52, 2003. (C) 2003 Wiley-Liss, Inc.

Genetic divergence between Cyprinus carpio carpio and Cyprinus carpio haematopterus as assessed by mitochondrial DNA analysis, with emphasis on origin of European domestic carp

Although common carp is the major fish species in Asian and European aquaculture and many domestic varieties have occurred, there is a controversy about the origination of European domestic common carp. Some scientists affirmed that the ancestor of European domestic common carp was Danube River wild common carp, but others considered it might be Asian common carp. For elucidating origination of European domestic common carp, we chose two representative European domestic common carp strains (German mirror carp and Russian scattered scaled mirror carp) and one wild common carp strain of Cyprinus carpio carpio subspecies (Volga River wild common carp) and two Asian common carp strains, the Yangtze River wild common carp (Cyprinus carpio haematopterus) and traditionally domestic Xingguo red common carp, as experimental materials. ND5-ND6 and D-loop segments of mitochondrial DNA were amplified by polymerase chain reaction and analyzed through restriction fragment length polymorphism (RFLP) and sequencing respectively. The results revealed that HaeIII and DdeI digestion patterns of ND5-ND6 segment and sequences of control region were different between European subspecies C. carpio carpio and Asian subspecies C. carpio haematopterus. Phylogenetic analysis showed that German mirror carp and Russian scattered scaled mirror carp belonged to two subspecies, C. carpio carpio and C. carpio haematopterus, respectively. Therefore, there were different ancestors for domestic carp in Europe: German mirror carp was domesticated from European subspecies C. carpio carpio and Russian scattered scaled mirror carp originated from Asian subspecies C. carpio haematopterus.

Reproduction mode of an artificial allotetraploid carp (Pisces; Cyprinidae)

Mature eggs of allotetraploid carp were activated by inactive sperm or crossed with normal sperms of common carp (Cyprinus carpio), crucian carp (Carassius auratus), Chinese blunt snout bream (Megalobrama amblycephala), Hemiculter leucisculus and Pseudorasbora parva. Chromosome counts showed that all offspring of these crosses presented a mode number of 200 chromosomes (4n = 200), and their morphological traits are much like maternal. Microsatelite marker and RAPD patterns between allotetraploid maternal and its offspring, reproduced from different paternal species, were identical. Cytological, morphological and molecular evidences suggested that allotetraploid carp female nucleus would not fuse with any male nucleus and its reproduction mode might be gynogenesis and therefore their offspring are retaining their tetraploidy and give origin to clonal individuals.

Introduction of the human lactoferrin gene into grass carp (Ctenopharyngodon idellus) to increase resistance against GCH virus

Haemorrhage can be an epidemic and fatal condition in grass carp. It is known now that the Grass Carp Haemorrhage Virus (GCHV) triggers haemorrhage. Human lactoferrin (hLF) plays an important role in the non-specific immune system, making some organisms more resistant to some viruses. Sperm of grass carp was mixed with linearized pCAhLFc, which is a DNA construct containing an hLF cDNA and the promoter of common carp beta-actin gene, and then electroporated. Then, mature eggs were fertilized in vitro with the treated sperm cells. The fry were sampled and analyzed by polymerase chain reaction (PCR). Results indicated that the foreign gene had been transferred successfully into the cells of some fry. Under optimal electroporation conditions, the efficiency of gene transfer was as high as 46.8%. About 35.7% of treated 5-month-old grass carp contained foreign genes. Most transgenic fry demonstrated significant delays in onset of symptoms of haemerrhage after injection of GCHV, suggesting a significant positive relationship between hLF cDNA and levels of disease resistance (P < 0.01). Results suggest that transgenic grass carp could be bred for increased resistance to haemorrhage. (C) 2002 Elsevier Science B.V. All rights reserved.

Growth and feed utilization in two strains of gibel carp, Carassius auratus gibelio: paternal effects in a gynogenetic fish

Gibel carp (Carassius auratus gibelio Bloch) is a natural gynogenetic fish which requires sperm of the same or related species to activate egg development. The eggs of one gibel carp were divided into two batches. One batch was 'fertilized' with sperm from gibel carp (strain DD), and the other 'fertilized' with sperm from red common carp (Cyprinus carpio red variety) (strain DR). The juveniles were transferred to the laboratory 36 days post-hatch. Triplicate groups of each strain were fed a formulated diet at either 3% or satiation ration for 8 weeks. At both the restricted and satiation rations, specific growth rate was significantly higher in strain DR than in strain DD. At the 3% ration, there was no significant difference in feeding rate or feed conversion efficiency between the two strains. At the satiation ration, strain DR had a significantly lower feeding rate but higher feed conversion efficiency than strain DD. At the satiation ration, strain DR had a significantly lower intake protein, but higher recovered protein than strain DD. There was no significant difference in faecal protein loss between the two strains. At the 3% ration, strain had no significant effects on intake protein, faecal protein or recovered protein. Neither faecal energy loss nor recovered energy was affected by strain or ration. At both the 3% and satiation ration, final body contents of dry matter and lipid were significantly lower in strain DR than strain DD, while there was no significant difference in protein and energy content between the two strains at either ration level. The results suggested that gibel carp 'fertilized' with sperm of common carp grew faster than those 'fertilized' with sperm of gibel carp through increased feed conversion efficiency and protein retention.

Differential screening and characterization analysis of the egg envelope glycoprotein ZP3 cDNAs between gynogenetic and gonochoristic crucian carp

Gynogenetic silver crucian carp, Carassius auratus gibelio, is an intriguing model. system. In the present work, a systemic study has been initiated by introducing suppression subtractive hybridization technique into this model system to identify the differentially expressed genes in oocytes between gynogenetic silver crucian carp and its closely related gonochoristic color crucian carp. Five differential cDNA fragments were identified from the preliminary screening, and two of them are ZP3 homologues. Moreover, the full length ZP3 cDNAs were cloned from their oocyte cDNA libraries. The length of ZP3 cDNAs were 1378 bp for gyno-carp and 1367 bp for gono-carp, and they can be translated into proteins with 435 amino acids. Obvious differences are not only in the composition of amino acids, but also in the number of potential O-linked oligosaccharide sites. In addition, gyno-carp ZP3 amino acid sequence has an unexpected higher identity value with common carp (83.5%) than that with the closely related gono-carp (74.7%). The unique homology may be originated from the ancient hybridization. Northern blot analysis confirmed that expression of the ZP3 gene occurred exclusively in the oocytes. Because O-linked oligosaccharides on ZP3 have been demonstrated to play very important roles in fertilization, it is suggested that the extra O-linked glycosylation sites may be related to the unique sperm-egg recognition mechanism in gynogenesis.

Genetic evidence for gonochoristic reproduction in gynogenetic silver crucian carp (Carassius auratus gibelio bloch) as revealed by RAPD assays

Sex evolution has been a debating focus in evolutionary genetics. In lower vertebrates of reptiles, amphibians, and fish, a species or a bioform reproduces either sexually or asexually but never both. A few species were found to consist of all females in fish. These all-female species can propagate by asexual reproduction modes, such as gynogenesis and hybridogenesis. However, the coexistence of sexuality and asexuality in a single species was recently noted only in a cyprinid fish silver crucian carp, Carassius auratus gibelio. This fish had been demonstrated to be capable of gynogenesis stimulated by sperm from other related species. Surprisingly, natural populations of this fish consist of a minor but significant portion (approx. 20%) of males. As different clones with specific phenotypic and genetic characteristics have been found, and RAPD markers specific to each clone have recently been identified, this fish offers many advantages for analyzing whether or not genetic recombination occurs between different clones. In this study, artificial propagation was performed in clone F and clone D. Ovulated eggs from clone F were divided into two parts and respectively inseminated with sperm from a clone D male and from a red common carp (Cyprinus carpio) male. The control clone D individuals were selected from gynogenetic offspring of clone D activated by sperm of red common carp. The phenotype and sex ratio in the experimental groups were also observed. Using RAPD molecular markers, which allow for reliable discrimination and genetic analysis of different clones, we have revealed direct molecular evidence for gonochoristic reproduction in the gynogenetic silver crucian carp and confirmed a previous hypothesis that the silver crucian carp might reproduce both gynogenetically and gonochoristically. Therefore, we conclude that the silver crucian carp possesses two reproductive modes, i.e., gynogenetic and gonochoristic reproduction. The response mechanism of two reproductive development modes may be the first discovery in vertebrates. Additionally, we discuss the evolutionary implication between gynogenetic and gonochoristic reproduction modes and the contribution of the minor proportion of males to genetic flexibility in the gynogenetic silver crucian carp.

Growth and feed utilization by F-4 human growth hormone transgenic carp fed diets with different protein levels

The F-4 generation of human growth hormone (hGH) transgenic red common carp Cyprinus carpio had significantly higher growth rates than the non-transgenic controls. Protein and energy intakes were significantly higher in the transgenic carp than in the controls fed the 20% protein diet, but were not different between the two strains fed diets with 30 and 40% protein. Faecal protein loss, as a proportion of protein intake, was significantly lower in the transgenics than in the controls fed diets with 20 and 30% protein, but was not different between the two strains Fed diet with 40% protein. Faecal energy loss, as a proportion of energy intake, was significantly lower in the transgenics than in the controls fed diet with 20% protein, but was not different between the two strains fed diets with 30 and 40% protein. Recovered protein, as a proportion of protein intake, was significantly higher in the transgenics than in the controls fed all diets, whereas recovered energy was significantly higher in the transgenic fish fed the 40% protein diet. For fish fed each diet, the transgenics had significantly higher body contents of dry matter and protein, but lower contents of lipid than the controls. It was concluded that transgenics were more efficient in utilizing dietary protein than the controls. it a lower dietary protein level; transgenics achieved higher growth rates mainly by increasing feed intake; at higher levels of dietary protein, transgenics achieved higher growth rates mainly through a higher energy conversion efficiency. (C) 1998 The Fisheries Society of the British Isles.

HORMONAL REPLACEMENT THERAPY IN FISH - HUMAN GROWTH-HORMONE GENE-FUNCTION IN HYPOPHYSECTOMIZED CARP

Transgenic common carp, Cyprinus carpio, produced by the microinjection of fertilized eggs with a linearized chimeric plasmid pMThGH, a human growth hormone (hGH) gene with a mouse metallothionein-I (MT) gene promoter in pBR322, were used to produce F1 and F2 transgenics. Following hypophysectomy of the transgenic F2 common carp, non-transgenic common carp and non-transgenic crucian carp, growth was monitored for up to 110 days. In addition, recombinant hGH was injected subcutaenously into a group of the non-transgenic crucian carp. Growth rate analyses indicated that (1) hypophysectomy of non-transgenic common carp and crucian carp results in the cessation of growth, (2) hGH administration can stimulate the growth of hypophysectomized crucian carp and (3) hypophysectomized hGH-transgenic common carp continue to grow in the absence of their own growth hormone, suggesting that the hGH-transgene is being expressed in tissues other than the pituitary.

1998 warmwater fish survey of Vancouver Lake, Clark County

The Vancouver Lake warmwater fish population was sampled during the fall of 1998. Vancouver Lake is a large lake, connected directly to the Columbia River through the Lake River to the north and the constructed flushing channel on the eastern shore. A total of 16 species of fish were encountered, mostly warmwater game fish with a few seasonal visitors. The lake has suffered from water quality problems, and during the 1970s, it was dredged, and a channel was created to aid in increasing the water turnover rate, effectively connecting both ends of the lake to the Columbia River. As there is direct access to the Columbia River, managing this lake as a closed system may prove difficult. It is of our opinion that it may be a more effective use of resources to increase the anglers access to the available resources through providing improved boat launch access, and fish habitat structure throughout the lake. Additionally, educating the public about the how-to's of common carp angling could possibly open up additional resources not commonly utilized by the public. (Document pdf contains 38 pages)

STREAM Journal, Vol. 6, No. 1, pp 1-21. Jan-March 2007

CONTENTS: Approaches to understanding pond-dike systems in Asia: the POND-LIVE project approach, by Dave Little, Marc Verdegem and Roel Bosma. The contribution of fish ponds to nutrient cycling in integrated farming systems, by P.N. Muendo, J.J. Stoorvogel and Marc Verdegem. Improving the contribution of fishfarming to livelihoods in Northeast Thailand, by Chittra Arjinkit, Roel Bosma, Danai Turongrouang. Benefits of pond-dike systems in Bangladesh, by M.S. Kabir, M.A. Wahab and Marc Verdegem. Common carp increases rohu production in farmers ponds, by Mohammad Mustafizur Rahman, Md. Abdul Wahab and Marc C.J. Verdegem. Improving pond-dike farming systems in the Mekong delta, Vietnam; the Can Tho approach, by Dang Kieu Nhan, Le Thanh Duong, Le Thanh Phong, Roel H. Bosma and Marc C.J. Verdegem. Fuzzy pathways for farm development in Vietnam, by Roel H. Bosma, Le Thanh Phong, and Dang Kieu Nhan.

Preliminary investigation on growth responses of Cyprinus carpio L. fed on locally formulated artificial diets

The growth rate and feed conversion ratios of the common carp, Cyprinus carpio were measured for five test diets in 14-day replicate laboratory studies. The young carp were fed with artificial test diets with crude protein contents ranging from 14.50 to 21.42 per cent. Within this range of feed characteristic optimum growth rates were obtained with diets containing 20.25 and 21.42 per cent crude protein. The study of the effect of varying ration levels showed that growth rates increased with increases of ration size, but the food conversion efficiency and protein efficiency ratios decreased markedly as ration size was increased

A review of recent advances in commercial tilapia culture

The paper introduces the tilapias as important culture fish; their classification, breeding habits and general ecology are discussed. The versatility and plasticity of their reproduction and growth are major advantages as well as draw backs in their use as culture species. The culture of tilapias in ponds, cages, pens and raceways are described and a comparison of extensive and intensive cultivation is made. The major factors in commercial cultivation of tilapias such as feeds, hatchery/fry production and disease control are discussed. Recommendation is made for the consideration of tilapia as the equivalent of the common carp in tropical African fish culture in either monoculture or in polyculture with other fish species

Aquaculture research: technology transfer for rural development in Nigeria

Conventional aquaculture has been promoted in Nigeria for the past five decades with minimal impact on rural communities: from the findings of Maclearen (1949) where he popularized the use of culturable fish predators Lutjanus sp; Pomades sp; Tarpon adanticus; Chrysichthys nigrodigitatus in earthen ponds near Onikan-Lagos, Nigeria; to the finding of Zwilling, 1963, who reported common carp, Cyprinus carpio propagation and culture in Panyan Fish Farm, near Jos; to the findings of FAO, 1965, when the potential culture of marine mullets culture in brackish water ponds in Buguma, Rivers State was presented. The work of other researchers Sivalingam, (1970; 1973), Ezenwa (1976), development officers and extension officers contributed to the development of aquaculture in few rural areas of the country and informed on public and private owned fish farm infrastructures. Despite a moderate long history of aquaculture research and development in Nigeria, an annual production level of 25,000 metric tons was recorded in 1999. This situation calls for a more sustainable approach for a stronger link between aquaculture research and technology transfer for the development of rural communities of Nigeria. This paper therefore examines some of the issues involved in the continuous flow of the new aquaculture technology in the improvement of fish protein output, standard of living of rural farmers and prevention of urban migration by the youth

Mono- and polyculture of silver barb (Puntius gonionotus) in deepwater rice systems in Bangladesh

Experiments with fish enclosures were conducted at the Deepwater Rice Farming Systems Research Site at Shuvullah, Mirzapur, Bangladesh. The objective was to study the performance of silver barb (Puntius gonionotus) called Thai sharputi or rajputi in Bangladesh in mono-and-polyculture with grass carp (Ctenopharyngodon idella), common carp (Cyprinus carpio), catla (Catla catla) and rohu (Labeo rohita). Each enclosure measured 21 m x 21 m with an approximate net height of 3.5 m. The stocking densities per cubic meter were 1 fingerling for Thai sharputi monoculture (enclosure 1), and 2 fingerlings for the polyculture systems (enclosure 2 and 3). The species ratio for enclosure 2 was 0.37:0.27:0.02:0.34 (grass carp:Thai sharputi:common carp:catla) and for enclosure 3, 0.4:0.4:0.2 (catla:rohu:Thai sharputi). In monoculture (enclosure 1), Thai sharputi performed well. This relatively good production was mainly attributed to the use of appropriately sized fingerlings and rapid growth from consumption of an abundant supply of azolla in addition to feed given. For the polyculture in enclosure 2, the average weight gain of common carp was the highest (673 g) followed by grass carp (475 g) and Thai sharputi (286 g). For the polyculture in enclosure 3, the length and weight gains for Thai sharputi were almost the same as for the monoculture.