Pterophyllum Research Articles

A new fish cell line derived from the caudal fin of freshwater angelfish Pterophyllum scalare: development and characterization

In this study, a new cell line derived from the caudal fin of the freshwater angelfish Pterophyllum scalare was developed and characterized. The cell line was designated angelfish fin (AFF) and subcultured 44 times since its development. These cells grew well in Leibovitz's -15 medium supplemented with 10% foetal bovine saline (FBS) at 28° C and the modal chromosome number (2n) was 48. The AFF cell-line is mainly comprised of epithelial cells as confirmed by immunocytological technique using anti-cytokeratin antibodies, an epithelial cell marker. This cell line was tested for growth in a temperatures range from 20 to 37° C and at various FBS concentrations of 5-20% at 28° C. The cell line was cryopreserved at different passage levels and revived successfully with 80% survival rate. Polymerase chain reaction amplification and sequencing of partial mitochondrial 16s rRNA and coI genes confirmed that the AFF cell-line originated from angelfish. Mycoplasma sp. contamination was not detected in AFF cells and checked by Hoechst 33258 fluorescence staining. At the 42nd passage the cells were transfected with 2 μg of pAcGFP1-N1 expression vector. The AFF cells exhibited cytotoxic effects when exposed to the bacterial extra cellular products from Serratia marcescens and Proteus hauseri. The AFF cells and cells from kidney and brain did not show cytopathic effect when exposed to cyprinid herpes virus2 and viral nervous necrosis virus. The newly developed AFF cell line will be useful for the isolation of viruses affecting angelfishes, such as iridoviruses, in the future.

The redox-active drug metronidazole and thiol-depleting garlic compounds act synergistically in the protist parasite Spironucleus vortens

Spironucleus vortens is a protozoan parasite associated with significant mortalities in the freshwater angelfish, Pterophyllum scalare. Control of this parasite is especially problematic due to restrictions on the use of the drug of choice, metronidazole (MTZ), on fish farms. Use of garlic (Allium sativum) is undergoing a renaissance following experimental validations of its antimicrobial efficiency. Ajoene ((E,Z)-4,5,9-trithiadodeca-1,6,11-triene 9-oxide), is a stable transformation product of allicin, the primary biologically active component of garlic. In the current study, an ajoene oil crude extract had a minimum inhibitory concentration (MIC) of 40μg/ml against S. vortens. GC–MS and NMR spectroscopy revealed this ajoene extract contained a mixture of the (E) and (Z)-ajoene isomers along with diallyl disulphide (DADS) and diallyl trisulphide (DATS). The only component of the ajoene crude oil found to substantially inhibit S. vortens growth by optical density monitoring (Bioscreen C Reader) was (Z)-ajoene (MIC 16μg/ml). Ajoene oil acted in synergy with MTZ in vitro, reducing the individual MIC of this drug (4μg/ml) by 16-fold, and that of ajoene oil by 200-fold with a fractional inhibitory concentration (FIC) index of 0.263. This synergistic interaction was confirmed in vivo. S. vortens-infected Pterophyllum scalare angelfish dosed orally with 0.5% (v/w) MTZ combined with 0.05% (v/w) ajoene displayed a significant reduction in faecal trophozoite count, whilst those fed on 0.5% MTZ flakes (half the recommended oral dose) alone did not. This study demonstrates for the first time the synergistic interaction between the synthetic drug MTZ and natural ajoene oil both in vitro and in vivo. Future work should evaluate the potential synergy of ajoene and MTZ against MTZ-resistant bacteria and protists.

Mycobacterium angelicum sp. nov., a non-chromogenic, slow-growing species isolated from fish and related to Mycobacterium szulgai.

The name 'Mycobacterium angelicum' dates back to 2003 when it was suggested for a slowly growing mycobacterium isolated from freshwater angelfish. This name is revived here and the novel species is proposed on the basis of the polyphasic characterization of four strains including the original one. The four strains presented 100 % 16S rRNA gene sequence similarity with Mycobacterium szulgai but clearly differed from M. szulgai for the milky white aspect of the colonies. The sequence similarity with the type strain of M. szulgai ranged, in eight additionally investigated genetic targets, from 78.9 to 94.3 %, an evident contrast with the close relatedness that emerged at the level of 16S rRNA gene. The average nucleotide identity between the genomes of M. szulgai DSM 44166T and strain 126/5/03T (type strain of the novel species) was 92.92 %, and supported the status of independent species. The confirmation of the name Mycobacterium angelicum sp. nov. is proposed, with strain 126/5/03T ( = CIP 109313T = DSM 45057T) as the type strain.

Spontaneous multi-cystic peripheral ameloblastoma in the freshwater angelfish, from the Brazilian state of Pará

Este trabalho registra a ocorrencia espontânea de ameloblastoma em P. scalare. O tumor foi obtido a partir de um exemplar de Acara Bandeira, sendo fixado, seccionado, e os fragmentos processados para microscopia de luz e microscopia eletronica de varredura (MEV). O exame macroscopico evidenciou uma massa tumoral que se estendia do rebordo alveolar do maxilar superior a face externa labial. A MEV, a neoplasia apresentou uma serie de espiculas. Microscopicamente, percebia-se um processo neoplasico constituido por tecido conjuntivo de caracteristica mixoide, ricamente vascularizado, onde eram observados fragmentos de tecido osteoide. A amostra revelou proliferacao neoplasica do epitelio odontogenico, onde as celulas neoplasicas se organizavam na forma de palicada. Alteracoes histopatologicas em peixes tem sido uteis biomarcadores do efeito a exposicao a substâncias toxicas, sendo as neoplasias lesoes especificas, comumente encontradas em peixes de areas poluidas, revelando uma associacao entre as lesoes e a exposicao a irritantes.

Monogenoidea on Exotic Indian Freshwater Fish. 3. Are Indian Guidelines for Importation of Exotic Aquarium Fish Useful and can they be Implemented; The Case of Neotropical Gussevia spiralocirra Kohn and Paperna, 1964

Gussevia spiralocirra , a Neotropical parasitic monogenoid (Platyhelminthes), has been recorded from the type host, freshwater angelfish Pterophyllum scalare (Cichlidae), collected from the post-quarantine populations in local aquarium markets of Lucknow and Itanagar, India. The finding establishes India as a geographical distribution record for G. spiralocirra , and more importantly, reveals a potentially serious breach of quarantine regulations of the country. The present communication provides a summary assessment of existing Indian guidelines for importation of exotic aquarium fish and highlights some of its major shortcomings.

Effect of Replacing Live Food with Formulated Feed on Reproductive Performance of Freshwater Angelfish, Pterophyllum scalare (Shcultze, 1823)

The combined effect of live food (LF) and formulated feed (FF) on reproductive performance of Pterophyllum scalare was studied. Mosquito larvae were used as live food in addition to formulated feed which contained 50% protein and 12% lipid. The experiment was arranged as a completely randomized design (CRD) with six replicates for each treatment: P. scalare were fed with formulated feed and/or live food: T1 (100% FF), T2 (75% FF and 25% LF), T3 (50% FF and 50% LF), T4 (25% FF and 75% LF) and T5 (100% LF). Relative fecundity, egg and oil globule diameter was significantly higher (P < 0.05) in T4 and T5 diets whereas inter spawning interval was lower in T3, T4, and T5 diets. Similarly hatching percentages were significantly higher with diets T3, T4, and T5. There were no significant differences observed between treatments on relative hatchling number, hatchling length, and weight. No significant difference was observed for egg and whole body composition of P. scalare fed with different diets. However there was a significant difference between treatments on fatty acid profile. It was concluded that live food can be replaced by up to 25-50% formulated feed without compromising reproductive performance of P. scalare

Non-invasive investigation of Spironucleus vortens transmission in freshwater angelfish Pterophyllum scalare

Spironucleus vortens is a protozoan fish parasite of veterinary and economic importance in the ornamental aquaculture industry. Despite this, key aspects of the life cycle of this organism, including its mode of transmission, have not been fully elucidated. We developed a non-invasive method for quantifying S. vortens in freshwater angelfish, which was then used to investigate parasite transmission and aggregation within host populations. As previously observed for S. meleagridis and S. salmonis, motile S. vortens trophozoites were detected in host faeces using light microscopy. Species-level identification of these flagellates was confirmed using 16S rDNA PCR. Faecal trophozoite counts were significantly correlated with trophozoite counts from the posterior intestine, the primary habitat of the parasite. This novel finding allowed effective prediction of intestinal parasite load from faecal counts. Overall, faecal count data revealed that 20% of hosts harbour 83% of parasites, conforming to the Pareto Principle (80/20 rule) of parasite aggregation with implications for parasite transmission. Trophozoites survived for ≥36 d outside the host within faeces and remained motile at low pH (comparable with that of angelfish stomach). No putative S. vortens cysts were observed in cultures or faecal samples. This calls into question the commonly accepted hypothesis that a protective cyst is required in the life cycle of S. vortens to facilitate transmission to a new host.

Effect of Clove Oil and Benzocaine on the Respiratory Metabolism of Angel Fish Pterophyllum Scalare

Anesthetics play an important role in fisheries research and aquaculture and are used to facilitate various handling procedures, such as weighing, sorting, collection of eggs and milt, tagging and medical treatment. Clove oil and benzocaine are approved anesthetic by US Food and Drug Administration. The present study investigates the effect of anesthetics on the respiratory metabolic parameters such as O 2 consumption rate, CO 2 output rate, NH 3 -N excretion rate, RQ, AQ and opercular activity of Angel fish, Pterophyllum scalare . The concentration of clove oil and benzocaine were selected based on the time of induction of sedation and recovery of the Angel fish. The concentration of clove oil selected for the Angel fish were 5 ppm, 15 ppm, 30 ppm, 45 ppm, 55 ppm and the benzocaine concentration were 30 ppm, 45 ppm, 50 ppm, 100 ppm, 200 ppm, 300 ppm and 400 ppm. The minimum values of respiratory metabolic parameters in clove oil-anaesthetized Angel fish P.scalare were observed at the concentration of 45 ppm while the same was in 50 ppm in the case of benzocaine treatment. Simulated transportation experiments were conducted for Angel fish and the results suggested that 50ppm benzocaine was the optimum dose of anaesthetics for safe transport of Angel fish.

Effects of Synbiotic (Biomin Imbo) on Growth Performance, Survival Rate, Reproductive Parameters of Angelfish (Pterophyllum scalare)

This study was carried out to evaluate the effect of synbiotic ( Biomin imbo ) on growth performance, survival rate and reproductive parameters of Angelfish ( Pterophyllum scalare ) via supplementation with Biomar. Four levels of Biomar experimental diets (54 % protein and 14 % lipid) were prepared by adding synbiotic (0.15, 0.5, 0.75, 1 g/Kg) at a basal diet (Biomar) and the Angelfish larvae in experimental treatments were fed four levels of synbiotic with 5 percent body weight (3 times a day). The larvae in the control treatment were fed without supplemented Biomar. The results showed that larvae fed on the synbiotic had significantly increased final body weight in comparison with the control treatment. After 90 days, 5 adult female and male Angelfish were divided from each treatment. The results showed that fecundity in experimental treatments increased in comparison with the control treatment. The synbiotic also had positive effects on hatching rate in comparison with those in the control treatment but, there were no significant differences (P > 0.05) among treatments. The synbiotic also had significant positive effects on the specific growth rate (SGR) and feed conversation efficiency (FCE) in comparison to those fed the control treatment, however treatment T3 was more than T4 (T3 > T4 > T2 > T1 > control treatment).

Polyculture of Freshwater AngelfishPterophyllum scalareand Pacific white shrimpLitopenaeus vannameiin low-salinity water

In this study, we evaluated the production potential of the polyculture of Freshwater Angelfish (Pterophyllum scalare, Cichlidae) and Pacific white shrimp (Litopenaeus vannamei). Two experimental trials were set up. The first trial entailed the use of a randomized design to investigate three treatments options: angelfish monoculture, shrimp monoculture, and fish and shrimp polyculture in 12 experimental 15 m2 ponds. In the second trial, we investigated two treatments (polyculture of caged fish and uncaged fish) in 10 experimental units. In trial 1, polyculture negatively affected fish growth and survival. However, fish did not affect shrimp growth and the greatest profit was achieved in polyculture. In trial 2, caging fish improve the growth of both fish and shrimp. An economic analysis showed the greatest profit and benefit cost ratio for caged fish compared with uncaged fish. We conclude that growing P. scalare and L. vannamei together is a better strategy than shrimp monoculture in low-salinity water. The use of caged fish in such a polyculture operation would enhance productivity and profitability.

Feeding level and frequency for freshwater angelfish

The objective of this study was to determine the optimal feeding level and feeding frequency for the culture of freshwater angelfish (Pterophyllum scalare). A randomized block design in a factorial scheme (3 × 2) with three feeding levels (30, 60 and 90 g/kg of body weight (BW)/day) and two feeding frequencies (1x and 2x/day) was set up in duplicate, representing 24 experimental units. Data were analyzed using two-way ANOVA and the Tukey test for comparison between means. After 84 days, results indicated that both factors influenced fish performance. No interaction between these factors was, however, observed. Increased feeding level and feeding frequency resulted in increased feed intake. The feed conversion ratio was negatively affected by feeding level, but not affected by feeding frequency. Final weights were higher when fish were fed twice daily, at levels of 60 or 90 g/kg BW/day. Specific growth rate was higher when fish received 60 or 90 g/kg BW/day, regardless of the feeding frequency. Survival was not affected by any treatment, with mean survival rates higher than 90%. It is recommended that juveniles be fed at a level of 60 g/kg BW/day with a minimum of two meals per day, to attain optimal survival, growth and feed efficiency.

Space size relative to prey width and total cover in an area both influence the habitat choices of freshwater angelfish Pterophyllum scalare in mesocosms

Prey survivorship can vary with average space size/prey width (Sp/Py) and total cover within an area (Ct/At). Hypotheses predict that prey survivorship is maximized at intermediate Sp/Py values and that prey survivorship increases rapidly from zero to low cover but increases at a slower rate as cover increases further. Freshwater angelfish Pterophyllum scalare made choices between artificial vegetation treatments consistent with these predictions. At constant low cover, fish did not exhibit any space width preferences. At constant medium cover, fish significantly preferred a narrower space treatment over a wider space treatment. At constant high cover, fish significantly preferred a wider space treatment over narrower space treatments. At constant Sp/Py, fish exhibited a marked preference for a higher-cover treatment at low-cover levels. They also preferred higher cover treatments under higher cover levels, but these preferences were not as pronounced. Sp/Py and Ct/At both influenced the fishes’ habitat choices.

Effect of Dietary Protein and Lipid Levels on Growth and Nutrient Utilization of Freshwater Angelfish Pterophyllum scalare

Nine semi-purified diets containing three levels of protein (300, 350, 400 g/kg) and three levels of lipid (60, 80, 100 g/kg) were fed ad libitum to juvenile freshwater angelfish Pterophyllum scalare (1.64±0.01 g) to determine the optimum dietary protein and lipid levels. Fish were stocked in 27 flow-through fiber-reinforced plastic tanks with 100 l water (10 fish/replicate) and fed 60 days. Fish were batch-weighed every 15 days to determine growth. The dietary protein level had a significant effect on protein efficiency rate (PER) but not on weight gain, feed conversion ratio (FCR), or specific growth rate (SGR). In contrast, the dietary lipid level had a significant effect on weight gain, FCR, and SGR but not on PER. The interaction of dietary protein and lipid had a significant effect on weight gain, FCR, SGR, and PER. No protein-sparing effect of the lipid was observed. Fish fed the diet containing 300 g protein and 60 g lipid per kg diet had a significantly better weight gain, SGR, PER, and FCR.

Vegetable oil sources in diets for freshwater angelfish (Pterophyllum scalare, Cichlidae): growth and thermal tolerance

The influence of fatty acid composition of the diets on the productive performance and on cold and heat tolerance of juvenile freshwater angelfish (Pterophyllum scalare), in three different phases, was studied. Phase I studied the productive performance of freshwater angelfish in a completely randomized experimental design with four treatments, canola, linseed, olive and soybean oils and four replicates during 50 days using 192 fish in 16 aquaria. Phase II studied the cold tolerance of juvenile freshwater angelfish using 72 juvenile freshwater angelfish, coming from phase I and maintained in 12 aquaria climatized chamber. The temperature was reduced 1ºC per day, until the observation of 100% fish mortality. Phase III, it was studied the heat tolerance of juvenile freshwater angelfish employing an identical procedure to phase II, but with a daily increase of 1ºC. Significant differences (P>0.05) were not observed for any parameters evaluated. Thus, it was concluded that the type of vegetable oil (canola, linseed, olive and soybean) used as a diet supplement did not affect the productive performance, nor the tolerance to cold and heat, of juvenile freshwater angelfish.

Anti-predatory behaviour of wild-caught vs captive-bred freshwater angelfish, Pterophyllum scalare

Summary Environments and experiences encountered in early life stages of animals shape their adult behaviour. When environments are maintained for several generations, differential selection forces act upon individuals to select those most fit to the particular conditions. As such, differences in the behaviour of captive bred and wild caught individuals have been observed recurrently. In fish, hatchery raised individuals tend to seek refuge less, making them more vulnerable to predators. We tested the hypothesis that captive breeding induces non-adaptive changes in behaviour of freshwater angelfish, Pterophyllum scalare. Wild-caught and captive-bred fish were exposed to a natural predator and measured for their anti-predator behaviours; no differences were found in behaviour under control conditions. When exposed to a natural predator, wild-caught fish exhibited significantly shorter freezing durations than captive-bred fish, and took significantly shorter time to resume normal behaviour. No differences in the time taken to initiate investigations of the predator were detected. The results demonstrate that captive-bred fish respond differently than their wild counterparts when exposed to a natural predator, and that this domestication has implications for captive rearing programmes.

Histochemical study on the intestine goblet cells in cichlid and poecilid species (Teleostei)

Histochemical properties of intestine goblet cells in firemouth cichlid, zebra mbuna, freshwater angelfish and platyfish are described. Goblet cells occurred regularly in the epithelial cell layer throughout the entire intestine, they were strongly coloured by alcian blue at pH 2.5. This colour got gradually weaker when the pH was reduced, but still after alcian blue at pH 0.2 these cells displayed a distinct blue colour. When the goblet cells were treated with periodic acid-Schiff (PAS), they displayed a strong purple-magenta colour. The findings that a number of goblet cells displayed various colours between blue and purple-magenta when acidic alcian blue was followed by PAS, and between blue and red-brown when acidic alcian blue was followed by neutral red, may reflect different ages or stages of development and differentiation for these cells. However, such results may also suggest a true cellular heterogeneity in the present population of goblet cells, reflecting that the intestine mucus layer has a number of roles in teleosts like lubrication, protection, immunological defence, digestion and absorption. In the ferritin injected specimens of firemouth cichlid and platyfish, a number of macrophage-like cells in intestine wall displayed Prussian blue precipitations in tissue treated with acid ferrocyanide, suggesting that these cells play a cleansing role in the intestinal wall. No ferritin uptake was seen in the intestine goblet cells and eosinophilic granule cells.

Gastric Cryptosporidiosis in Freshwater Angelfish (Pterophyllum Scalare)

A freshwater angelfish (Pterophyllum scalare) hatchery experienced variable levels of emaciation, poor growth rates, swollen coelomic cavities, anorexia, listlessness, and increased mortality within their fish. Multiple chemotherapeutic trials had been attempted without success. In affected fish, large numbers of protozoa were identified both histologically and ultrastructurally associated with the gastric mucosa. The youngest cohort of parasitized fish was the most severely affected and demonstrated the greatest morbidity and mortality. The protozoa were morphologically most consistent with Cryptosporidium. All of the protozoan life stages were identified ultrastructurally and protozoal genomic DNA was isolated from parasitized tissue viscera and sequenced. Histological, ultrastructural, genetic, and phylogenetic analyses confirmed this protozoal organism to be a novel species of Cryptosporidium.

Dietary protein and energy requirements of juvenile freshwater angelfish

Dietary protein and energy requirements of juvenile freshwater angelfish (Pterophyllum scalare) were evaluated. A 3 × 2 factorial design was used, with three dietary crude protein levels being tested (26, 30, and 34% of CP) combined with two digestible energy levels (3,100 and 3,300 kcal DE/kg of diet) in three replicates. Juveniles averaging 2.33 ± 0.26 g were reared in a 25L-aquarium with controlled temperature (26 ± 1ºC), biological filter and stocking density of six fish/aquarium. Fish were fed ad libitum at 09:00 a.m., 2:00 p.m. and 4:30 p.m. The following performance parameters were evaluated: final weight, final length, weight gain, feed intake, feed conversion ratio, specific growth rate, protein efficiency ratio and condition factor. Fish fed diets with 26% CP showed greater protein efficiency values when compared to those fed diets with 34% CP. Diets with 26% of CP and 3100 kcal DE/kg could meet the nutritional requirements of juvenile freshwater angelfish.

Effects of commonly used disinfectants and temperature on swim bladder non-inflation in freshwater angelfish, Pterophyllum scalare (Lichtenstein)

Swim bladder non-inflation is common in hatchery-reared fish and was also reported in angelfish, Pterophyllum scalare L. Fish eggs are routinely treated with disinfectants and exposed to a variety of physical conditions. We studied the effect of these factors on the prevalence of swim bladder non-inflation in angelfish. Hatching and rearing at 26 °C resulted in 8% swim bladder non-inflation, compared to no swim bladder non-inflation at 28 °C. Eggs hatched in the presence of 1, 2 and 5 ppm methylene blue exhibited significant increases in swim bladder non-inflation (11, 9 and 33%, respectively; none in controls). Time of exposure to methylene blue was a key factor. Exposure for up to 1 day post-hatch did not affect swim bladder non-inflation, but exposure from 2 days onwards significantly increased swim bladder non-inflation. Hydrogen peroxide at 250 ppm significantly increased swim bladder non-inflation (65% comparing to 27% in the control). Higher concentrations resulted in 100% mortality. Exposure to acriflavin at 2.25 ppm, but not 1.25 ppm, significantly increased swim bladder non-inflation (75 and 52% respectively; 20% in controls). Chloramine-T did not significantly affect swim bladder non-inflation. Treatment with methylene blue and acriflavin, but not chloramine-T, produced significantly different frequencies of swim bladder non-inflation morphologies compared to control fish. In conclusion, several chemical treatments commonly used in aquaculture and sub-optimal water temperature increase the prevalence of swim bladder non-inflation in angelfish.

Sistemas de criação para o acará-bandeira (<em>Pterophyllum scalare</em>)

The present study aimed to compare three production systems for freshwater angelfish: intensive indoor aquarium, semi-intensive greenhoused earth pond (monoculture) and greenhoused earth pond in polyculture with Amazon River prawn. Two stock densities were also tested: 1.6 and 3.2 fish 10 L -1 . The experimental design was entirely randomized, in a 3x2 factorial scheme, with four replicates. ANOVA and Tukey’s test were used (α = 0.05). Fish from monoculture and polyculture systems showed over twice the weight gain, lower food consumption, better food conversion rate, bigger standard length and group uniformity than the fish maintained in aquariums. The aquarium system showed lower survival than monoculture. The stock density increase reduced the weight gain, standard length and body condition in all culture systems. The interaction analysis for biomass was statistical significance, showing no difference between the densities inside aquarium and increasing with density within mono and polyculture ponds. In polyculture, mean final weight and survival of Amazon river prawns were 1.66 g e 84.4%, respectively. Due to better growth results, density of 1.6 fish 10 L -1 is better than 3.2 fish 10 L -1 . Freshwater angelfish can be produced better in hapas on monocultured or polycultured semi-intensive compared to indoor aquariums system.